Human anti-B7RP1 Neutralizing Antibodies

ABSTRACT

This invention provides antibodies that interact with or bind to human B7 related protein-1 (B7RP1) and antibodies that bind to and neutralize the function of B7RP1 thereby. The invention also provides pharmaceutical compositions of said antibodies and methods for neutralizing B7RP1 function, and particularly for treating immune disorders (e.g., inappropriate immune response) by administering a pharmaceutically effective amount of anti-B7RP1 antibodies. Methods of detecting the amount of B7RP1 in a sample using anti-B7RP1 antibodies are also provided.

This application claims priority to U.S. provisional patent applicationSer. No. 60/700,265, filed Jul. 18, 2005, the disclosure of which isexplicitly incorporated by reference herein.

FIELD OF THE INVENTION

The invention relates to human monoclonal antibodies that bind B7related protein-1 (B7RP1). Compositions and methods for treatingdiseases and disorders related to immunosuppression and immuneactivation are also described.

BACKGROUND OF THE INVENTION

T-cells initiate the immune response, mediate antigen-specific effectorfunctions, and regulate the activity of other leukocytes by secretingcytokines. For the generation of a proper T-lymphocyte (T-cell) immuneresponse, two signals must be provided to the T-cell by antigenpresenting cells (APC). Antigen must be presented to the T-cell receptor(TCR) via a major histocompatibility complex (MHC), in an event thatdetermines specificity. TV-cells can only recognize antigen presented onan APC. In addition to the antigen receptor, proper T-cell activationalso requires the interaction of other cell-surface molecules on boththe T-cell and the APC. These molecules, referred to as co-stimulatorymolecules, consist of a receptor on the responding cell and a ligandpresent on the inducer cell. This antigen independent, co-stimulatorysignal must be delivered by engagement of members of the B7 family onthe APC with their receptors on T-cells. A productive immune responseleads to proliferation, differentiation, clonal expansion, and effectorfunction. In the absence of the second, co-stimulatory signal, T-cellsundergo a state of long-lasting antigen-specific unresponsiveness,termed anergy. Phase II clinical experiments have demonstrated thatblocking one co-stimulation pathway is efficacious in the treatment ofpsoriasis (Abrams et al., 2000, J Exp Med 192:681-94; Abrams et al.,1999, J. Clin. Invert, 103:1243-52) and rheumatoid arthritis (Kremer etal., 2003, New England Journal of Medicine 349:1907-15), indicating thatthis general strategy is a good target for immunomodulatory therapy.

A particular co-stimulatory B7 molecule, B7 related protein-1 (B7RP1),is a type 1 transmembrane protein with a signal sequence andextracellular domain at the amino-terminus, an extracellular domaincomprising two Ig loops, a transmembrane domain, and a carboxy terminalintracellular domain (PCT Application Publication No. WO 00/46240).B7RP1 preferentially binds to ICOS (which stands for “induciblecostimulator”; Yoshing a et al., 2000, Int Immun. 12:1439-1447)expressed on the cell surface of T-cells. ICOS plays an important rolein the production of both type 1 and type 2 cytokines by activatedT-cells (Coyle et al., 2000, Immunity 13:95-105).

B7RP1 is the sole ligand expressed constitutively on APCs (Yoshinaga etal., 1999, Nature. 402:827-32), while ICOS is expressed only onactivated T-cells (McAdam et al., 2000, Journal of Immunology165:5035-40). B7RP1-dependent signaling is required for the activationof the effector (i.e. fully activated) T-cell, as well as its maturationfrom its naïve precursor (Dong et al., 2003, Journal of Autoimmunity.21:255-60; Coyle et al., 2000, Immunity. 13:95-105). Consequently, theB7RP1/ICOS interaction is required for proper T-cell-dependent recallimmune responses (Dong et al., 2003, Journal of Autoimmunity.21:255-60).

Current attempts to interfere with the co-stimulatory T-cell pathwayhave focused primarily on co-stimulatory polypeptides that block T-cellactivation only, but have not focused on activation and maturation.Consequently, these therapies provide general inhibition of T-cellfunction. In contrast, blocking the B7RP1/ICOS interaction provides amore specific inhibition of T-cell function by affecting only matureeffector T-cells. Thus, blocking the B7RP1/ICOS interaction in aclinical setting is highly desirable because it would provide a morelimited side-effect profile than co-stimulation therapies that blocknaïve T-cell activation only.

SUMMARY OF THE INVENTION

The invention provides monoclonal antibodies that bind to B7 relatedprotein-1 (B7RP1). In one embodiment, the monoclonal antibodies arehuman monoclonal antibodies that neutralize biological activities ofB7RP1 and are particularly useful for inhibiting partially or completelythe immune co-stimulatory activity of B7RP1. Also provided by theinvention are cells, particularly hybridoma cells that produce themonoclonal antibodies of the invention. In particular aspects, theantibodies of the invention bind specifically to the H or D region ofB7RP1 as described herein.

The invention further provides fusion proteins comprising the sequenceof an antibody Fc region and one or more sequences identified as SEQ IDNO: 1 through SEQ ID NO. 40. Such molecules can be prepared usingmethods as described, for example, in International Patent Application,Publication No. WO 00/24782, which is hereby incorporated by reference.Such molecules can be expressed, for example, in mammalian cells (e.g.Chinese Hamster Ovary cells) or bacterial cells (e.g. E coli cells).

In certain aspects, the invention provides antibodies comprising a heavychain and a light chain, wherein the heavy chain comprises an heavychain constant region selected from IgG1, IgG2, IgG3, IgG4, IgM, IgA andIgE heavy chain constant regions or any allelic variation thereof (asdiscussed in Kabat et al., 1991, SEQUENCES OF PROTEINS OF IMMUNOLOGICALINTEREST, Fifth Edition, U.S. Department of Health and Human Services,NIH Publication No. 91-3242), incorporated herein by reference, and thevariable region of the heavy chain comprises an amino acid sequence asset forth in any of SEQ ID NO: 7 through SEQ ID NO. 14, or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof. An antibody of the invention comprises either an amino acidsequence of the IgG2 heavy chain constant region as set forth in SEQ IDNO: 41 or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof, or an amino acid sequence of the IgG1heavy chain constant region as set forth in SEQ ID NO: 42 or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof. In certain embodiments, the antibodies are monoclonalantibodies, human antibodies, or preferably human monoclonal antibodies.

In certain aspects, the invention provides antibodies comprising a heavychain and a light chain, wherein the light chain comprises a constantregion having an amino acid sequence as set forth in SEQ ID NO: 43 or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof, and the light chain variable region comprises an amino acidsequence as set forth in any of SEQ ID NO: 1 through SEQ ID NO, 6, or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof. In certain embodiments, the antibodies are monoclonalantibodies, human antibodies, or preferably human monoclonal antibodies.

In certain aspects, antibodies of the invention comprise a heavy chainand a light chain, wherein the variable region of the heavy chaincomprises an amino acid sequence as set forth in SEQ ID NO: 7 or SEQ IDNO: 8, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof. In other aspects, the light chainvariable region comprises an amino acid sequence as set faith in SEQ IDNO: 1, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof.

In other aspects, antibodies of the invention comprise a heavy chain anda light chain, wherein the variable region of the heavy chain comprisesan amino acid sequence as set forth in SEQ ID NO: 9, or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof. In other aspects, the light chain variable region comprises anamino acid sequence as set forth in SEQ ID NO: 2, or an antigen-bindingor an immunologically functional immunoglobulin fragment thereof.

In additional aspects, the heavy chain comprises at least onecomplementarity determining region (CDR) having an amino acid sequenceas set forth in any of SEQ ID NO: 27 through SEQ ID NO. 40, or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof. In still further aspects, the light chain comprises at leastone CDR having an amino acid sequence as set forth in any of SEQ ID NO:15 through SEQ ID NO. 26, or an antigen-binding or an immunologicallyfunctional immunoglobulin fragment thereof:

The invention also provides antibodies that bind specifically to B7RP1,wherein the heavy chain comprises a variable region comprising an aminoacid sequence as set forth in SEQ ID NO: 7 or SEQ ID NO: 8, or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof, and the light chain comprises a variable region comprising anamino acid sequence as set forth in SEQ ID NO: 1, or an antigen-bindingor an immunologically functional immunoglobulin fragment thereof.

In addition, the invention provides antibodies that bind specifically toB7RP1, wherein the heavy chain comprises a variable region comprising anamino acid sequence as set forth in SEQ ID NO: 9, or an antigen-bindingor an immunologically functional immunoglobulin fragment thereof, andthe light chain comprises a variable region comprising an amino acidsequence as set forth in SEQ ID NO: 2, or an antigen-binding or animmunologically functional immunoglobulin fragment thereof.

In certain aspects, the invention also provides antibodies, comprising aheavy chain and a light chain, wherein the heavy chain comprises a heavychain variable region, and wherein the heavy chain variable regioncomprises a sequence that has at least about 75%, at least about 80%, atleast about 85%, at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or at least about 99% identity to the amino acid sequence as setforth in any of SEQ ID NO: 7 through SEQ ID NO. 14, and wherein thelight chain comprises a light chain variable region, and wherein thelight chain variable region comprises a sequence that has at least about80%, at least about 85%, at least about 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or at least about 99% identity to the amino acid sequenceas set forth in any of SEQ ID NO: 1 through SEQ ID NO. 6, wherein theantibody binds specifically to B7RP1.

The invention also provides antibodies that bind specifically to B7RP1,wherein the heavy chain comprises an amino acid sequence as set forth inSEQ ID NO: 44 or SEQ ID NO: 46, or an antigen-binding or animmunologically functional immunoglobulin fragment thereof, and thelight chain comprises an amino acid sequence as set forth in SEQ ID NO:45, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof.

The invention also provides antibodies that bind specifically to B7RP1,wherein the heavy chain comprises an amino acid sequence as set forth inSEQ ID NO: 47, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof, and the light chain comprises an aminoacid sequence as set forth in SEQ ID NO: 48, or an antigen-binding or animmunologically functional immunoglobulin fragment thereof.

In certain aspects, the invention provides antibodies, comprising aheavy chain and a light chain, wherein the heavy chain comprises a heavychain variable region, and wherein the heavy chain variable regioncomprises at least one CDR having a sequence that has at least about75%, at least about 80%, at least about 85%, at least about 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least about 99% identity to theamino acid sequence as set forth in any of SEQ ID NO: 27 through SEQ IDNO. 40, and wherein the light chain comprises a light chain variableregion, and wherein the light chain variable region comprises at leastone CDR having an amino acid sequence that has as least about 80%, atleast about 85%, at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or at least about 99% identity to the amino acid sequence as setforth in SEQ ID NO: 15 through SEQ ID NO. 26, wherein the antibody bindsspecifically to B7RP1.

The invention also provides single chain antibodies, single chain Fvantibodies, F(ab) antibodies, F(ab)′ antibodies and (Fab′)₂ antibodies.

In particular aspects, the invention provides a light chain comprisingan amino acid sequence as set forth in SEQ ID NO: 15 through SEQ ID NO.26, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof.

In addition, the invention provides a heavy chain comprising an aminoacid sequence as set forth in any of SEQ ID NO: 27 through SEQ ID NO.40, or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof.

The invention also relates to isolated human antibodies thatspecifically bind B7RP1, wherein the antibody comprises: (a) human heavychain framework regions, a human heavy chain CDR1 region, a human heavychain CDR2 region, and a human heavy chain CDR3 region; and (b) humanlight chain framework regions, a human light chain CDR1 region, a humanlight chain CDR2 region, and a human light chain CDR3 region. In certainaspects, the human heavy chain CDR1 region can be the heavy chain CDR1region as shown in any of SEQ ID NO: 27, 30, or 35 and the human lightchain CDR1 region can be the light chain CDR1 region shown in any of SEQID NO: 15, 18, or 24. In other aspects, the human heavy chain CDR2region can be the heavy chain CDR2 region as shown in any of SEQ ID NO:28, 31, 33, 36, or 39, and the human light chain CDR2 region can be thelight chain CDR2 as shown in any of SEQ ID NO: 16, 19, or 21. In stillother aspects, the human heavy chain CDR3 region is the heavy chain CDR3region as shown in any of SEQ ID NO: 29, 32, 34, 37, 38 or 40, and thehuman light chain CDR3 region is the light chain CDR3 region as shown inany of SEQ ID NO: 17, 20, 22, 23, 25, or 26.

The antibodies of the invention are characterized by the ability to bindspecifically to B7RP1. Furthermore, antibodies of the invention have thecapacity to antagonize at least one in vitro and/or in vivo activityassociated with B7RP1 polypeptides. The invention provides isolatedanti-human B7RP1 human antibodies with high affinity binding to B7RP1polypeptides, wherein the antibodies bind to a human B7RP1 polypeptideand dissociates from the human B7RP1 polypeptide with a dissociationconstant (K_(D)) of about 10⁻⁶ M, 10⁻⁷ M, 10⁻⁸ M, 10⁻⁹ M, 10⁻¹⁰ M, 10⁻¹¹M, 10⁻¹² M, or less, as determined using KinExA, or which inhibit B7RP1induced survival in an in vitro neutralization assay with an EC₅₀ ofabout 10⁻⁶ M, 10⁻⁷ M, 10⁻⁸ M, 10⁻⁹M, 10⁻¹⁰ M, 10⁻¹¹M, 10⁻¹² M, or less.

The invention also provides isolated human antibodies or anantigen-binding or immunologically functional immunoglobulin fragmentsthereof that bind specifically to B7RP1, wherein the antibodies orfragments comprise a heavy chain variable region comprising a heavychain CDR1, CDR2, and CDR3, wherein:

-   -   a) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 27, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 28, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 29;    -   b) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 30, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 31, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 32;    -   c) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 27, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 33, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 34;    -   d) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 35, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 36, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 37;    -   e) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 27, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 33, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 38; or    -   f) the heavy chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 35, the heavy chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 39, and the heavy chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 40.

The invention also provides an isolated human antibody or anantigen-binding or an immunologically functional immunoglobulin fragmentthereof that binds specifically to B7RP1, wherein the antibody orfragment comprises a light chain variable region comprising a lightchain CDR1, CDR2, and CDR3, wherein:

-   -   a) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 15, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 16, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 17;    -   b) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 18, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 19, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 20;    -   c) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 15, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 21, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 22;    -   d) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 18, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 19, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 23;    -   e) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 24, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 16, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 25; or    -   f) the light chain CDR1 has an amino acid sequence as set forth        in SEQ ID NO: 24, the light chain CDR2 has an amino acid        sequence as set forth in SEQ ID NO: 16, and the light chain CDR        3 has an amino acid sequence as set forth in SEQ ID NO: 26.

The invention also provides antibodies that compete with binding of theantibodies described herein to B7RP1. In certain aspects, a competitiveantibody of the invention competes with binding of an antibody thatcomprises any of SEQ ID NO: 1-40 to human B7RP1

Also part of the invention are polynucleotide sequences that encodeanti-human B7RP1 human antibodies, vectors comprising the polynucleotidesequences encoding anti-human B7RP1 human antibodies, host cellstransformed with vectors incorporating polynucleotides that encodeanti-human B7RP1 human antibodies, formulations comprising anti-humanB7RP1 human antibodies and methods of making and using same.

The invention also provides methods for detecting B7RP1 in a biologicalsample, comprising the step of contacting the sample with an antibody ofthe invention or antigen-binding fragment thereof. An anti-B7RP1antibody of the invention may be employed in any known assay method,such as competitive binding assays, direct and indirect sandwich assays,immunoprecipitation assays and enzyme-linked immunosorbent assays(ELISA) (See, Sola, 1987, Monoclonal Antibodies: A Manual of Techniques,pp. 147-158, CRC Press, Inc.) for the detection and quantitation ofB7RP1. The antibodies can bind B7RP1 with an affinity that isappropriate for the assay method being employed.

In addition, the invention provides methods for treating a diseaseassociated with increased production of B7RP1, increased sensitivity toB7RP1, and/or diseases related to control of T-cell responses,comprising the step of administering a pharmaceutically effective amountof a pharmaceutical composition comprising at least one antibody of theinvention or an antigen-binding or an immunologically functionalimmunoglobulin fragment thereof to an individual in need thereof.

Embodiments of the invention will become evident from the followingdetailed description and the claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A depicts the 16H antibody variable region sequence (SEQ ID NO: 7)and the corresponding 16H variable region germline (16Hg) sequence (SEQID NO: 8).

FIG. 1B depicts results of co-stimulation assays using anti-CD3 andhB7RP1-Fc fusion protein demonstrating that 16Hg retains its biologicalactivities compared with 16H.

FIG. 2 shows the results of Biacore® binding assays with 16H, 16Hg, and5D antibodies.

FIG. 3 shows the results of KinExA binding assay with 5D antibody.

FIG. 4 shows the results of KinExA binding assay with 2H antibody.

FIG. 5 shows the results of KinExA binding assay with 2H germline (2Hg)antibody.

FIG. 6 depicts the results of binding-competition assays showing that16H antibody competes away binding of ICOS-Fc on B7RP-1, analyzed byflow cytometry.

FIG. 7 depicts a summary of a B7RP-1 single nucleotide polymorphism(SNP) analysis.

FIG. 8 depicts a summary of the analysis of a set of anti-human B7RP-1monoclonal antibodies in ELISA competition assays. Values shown areIC₅₀s for inhibition of binding of an ICOS-Fc fusion protein.

FIG. 9A shows fluorescent staining of B7RP1 extracellular domain (ECD)with labeled 16H, 5D, and ICOS antibodies.

FIG. 9B shows similar binding efficacy of 16H and 5D antibodies to aB7RP1 SNP variant.

FIG. 9C depicts the results of co-stimulation assays with 16H or 5Dantibodies and SNP variants.

FIG. 10A shows plate co-stimulation assay results with 1B7v2 monoclonalantibodies compared with a number of different anti-murine B7RP-1monoclonal antibodies.

FIGS. 10B, 10C, and 10D show the results of antigen challengeexperiments, analyzed for antigen-specific serum IgM (FIG. 10B), IgG2a(FIG. 10C), and IgG1 (FIG. 10D).

FIG. 11 depicts ELISA results demonstrating that serum IL-5 levels arerepressed by 1B7v2 antibodies.

FIG. 12A shows that 16H antibodies can bind to cynomolgus monkey B7RP1(right panel) and human B7RP1 (left panel).

FIG. 12B shows that 16H, 16Hg, and 5D antibodies can inhibit cynomolgusmonkey B7RP1/ICOS-dependent T cell activation.

FIG. 13A depicts individual cynomolgus monkey and group mean titervalues at day 53 and day 57 after secondary challenge with tetanustoxoid on day 42 in animals treated with 16H antibodies.

FIG. 13B depicts individual cynomolgus monkey and group mean titervalues at day 53 and day 57 after secondary challenge with tetanustoxoid on day 42 in animals treated with 5D antibodies.

DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS

The section headings used herein are for organizational purposes onlyand are not to be construed as limiting the subject matter described.All references cited in this application are expressly incorporated byreference herein for any purpose.

Definitions

Conventional techniques may be used for recombinant DNA, oligonucleotidesynthesis, and tissue culture and transformation (e.g., electroporation,lipofection). Enzymatic reactions and purification techniques may beperformed according to manufacturer's specifications or as commonlyaccomplished in the art or as described herein. The foregoing techniquesand procedures may be generally performed according to methods wellknown in the art and as described in various general and more specificreferences that are cited and discussed throughout the presentspecification. See e.g., Sambrook et al., 2001, MOLECULAR CLONING: ALABORATORY MANUAL, 3d ed., Cold Spring Harbor Laboratory Press, ColdSpring Harbor, N.Y., which is incorporated herein by reference for anypurpose. Unless specific definitions are provided, the nomenclatureutilized in connection with, and the laboratory procedures andtechniques of, analytical chemistry, synthetic organic chemistry, andmedicinal and pharmaceutical chemistry described herein are those wellknown and commonly used in the art. Similarly, conventional techniquesmay be used for chemical syntheses, chemical analyses, pharmaceuticalpreparation, formulation, and delivery, and treatment of patients.

As utilized in accordance with the present disclosure, the followingterms, unless otherwise indicated, shall be understood to have thefollowing meanings. The phrases “biological property”, “biologicalcharacteristic”, and the term “activity” in reference to an antibody ofthe present invention are used interchangeably herein and include, butare not limited to, epitope affinity and specificity (e.g., anti-humanB7RP1 human antibody binding to human B7RP1), ability to antagonize theactivity of the targeted polypeptide (e.g., B7RP1 activity), the in vivostability of the antibody, and the immunogenic properties of theantibody. Other identifiable biological properties or characteristics ofan antibody recognized in the art include, for example,cross-reactivity, (i.e., with non-human homologs of B7RP1, or with otherproteins or tissues, generally), and ability to preserve high expressionlevels of protein in mammalian cells. The aforementioned properties orcharacteristics can be observed or measured using art-recognizedtechniques including, but not limited to ELISA, competitive ELISA,surface plasmon resonance analysis, in vitro and in vivo neutralizationassays (e.g., Example 2), and immunohistochemistry with tissue sectionsfrom different sources including human, primate, or any otherappropriate source. Particular activities and biological properties ofanti-human B7RP1 human antibodies are described in further detail in theExamples below.

The term “isolated polynucleotide” as used herein shall mean apolynucleotide of genomic DNA, cDNA, RNA, or synthetic origin or somecombination thereof which by virtue of its origin the isolatedpolynucleotide (1) is not associated with all or a portion of apolynucleotide with which the isolated polynucleotide is found innature, (2) is linked to a polynucleotide to which it is not linked innature, or (3) does not occur in nature as part of a larger sequence.

The term “polynucleotide” as referred to herein means single-stranded ordouble-stranded nucleic acid polymers of at least 10 nucleotides inlength. In certain embodiments, the nucleotides comprising thepolynucleotide can be ribonucleotides or deoxyribonucleotides or amodified form of either type of nucleotide. Said modifications includebase modifications such as bromuridine, ribose modifications such asarabinoside and 2′,3′-dideoxyribose and internucleotide linkagemodifications such as phosphorothioate, phosphorodithioate,phosphoroselenoate, phosphorodiselenoate, phosphoroanilothioate,phoshoraniladate and phosphoroamidate. The term “polynucleotide”specifically includes single and double stranded forms of DNA or RNA.

The term “oligonucleotide” referred to herein includes naturallyoccurring, and modified nucleotides linked together by naturallyoccurring, and/or non-naturally occurring oligonucleotide linkages.Oligonucleotides are a polynucleotide subset comprising members that aregenerally single-stranded and have a length of 200 nucleotides or fewer.In certain embodiments, oligonucleotides are 10 to 60 nucleotides inlength. In certain embodiments, oligonucleotides are 12, 13, 14, 15, 16,17, 18, 19, or 20 to 40 nucleotides in length. Oligonucleotides may besingle stranded or double stranded, e.g. for use in the construction ofa genetic mutant. Oligonucleotides of the invention may be sense orantisense oligonucleotides with reference to a protein-coding sequence.

The term “naturally occurring nucleotides” includes deoxyribonucleotidesand ribonucleotides. The term “modified nucleotides” includesnucleotides with modified or substituted sugar groups and the like. Theterm “oligonucleotide linkages” includes oligonucleotide linkages suchas phosphorothioate, phosphorodithioate, phosphoroselenoate,phosphorodiselenoate, phosphoroanilothioate, phoshoraniladate,phosphoroamidate, and the like. See, e.g., LaPlanche et al., 1986, Nucl.Acids Res., 14:9081; Stec et al., 1984, J. Am. Chem. Sac., 106:6077;Stein et al., 1988, Nucl. Acids Res., 16:3209; Zon et al., 1991,Anti-Cancer Drug Design, 6:539; Zon et al., 1991, OLIGONUCLEOTIDES ANDANALOGUES: A PRACTICAL APPROACH, pp. 87-108 (E. Eckstein, Ed.), OxfordUniversity Press, Oxford England; Stec et al., U.S. Pat. No. 5,151,510;Uhlmann and Peyman, 1990, Chemical Reviews, 90:543, the disclosures ofwhich are hereby incorporated by reference for any purpose. Anoligonucleotide can include a detectable label to enable detection ofthe oligonucleotide or hybridization thereof.

The term “isolated protein” referred to herein means that a subjectprotein (1) is free of at least some other proteins with which it wouldbe found in nature, (2) is essentially free of other proteins from thesame source, e.g., from the same species, (3) is expressed by a cellfrom a different species, (4) has been separated from at least about 50percent of polynucleotides, lipids, carbohydrates, or other materialswith which it is associated in nature, (5) is not associated (bycovalent or noncovalent interaction) with portions of a protein withwhich the “isolated protein” is associated in nature, (6) is operablyassociated (by covalent or noncovalent interaction) with a polypeptidewith which it is not associated in nature, or (7) does not occur innature. Such an isolated protein can be encoded by genomic DNA, cDNA,mRNA or other RNA, of synthetic origin, or any combination thereof. Inone embodiment, the isolated protein is substantially free from proteinsor polypeptides or other contaminants that are found in its naturalenvironment that would interfere with its use (therapeutic, diagnostic,prophylactic, research or otherwise).

An “isolated” antibody is one that has been identified and separatedand/or recovered from a component of its natural environment.Contaminant components of its natural environment are materials thatwould interfere with diagnostic or therapeutic uses for the antibody,and may include enzymes, hormones, and other proteinaceous ornon-proteinaceous substances. In certain embodiments, the antibody ispurified (1) to greater than 95% or greater than 99% by weight ofantibody as determined by the Lowry method, (2) to a degree sufficientto obtain at least 15 residues of N-terminal or internal amino acidsequence by use of a spinning cup sequenator, or (3) to homogeneity bySDS-PAGE under reducing or nonreducing conditions using Coomassie blueor silver stain. Isolated antibody includes the antibody in situ withinrecombinant cells since at least one component of the antibody's naturalenvironment will not be present.

The terms “polypeptide” or “protein” means molecules having the sequenceof native proteins, that is, proteins produced by naturally-occurringand specifically non-recombinant cells, or genetically-engineered orrecombinant cells, and comprise molecules having the amino acid sequenceof the native protein, or molecules having deletions from, additions to,and/or substitutions of one or more amino acids of the native sequence.The terms “polypeptide” and “protein” specifically encompass anti-B7RP1antibodies, or sequences that have deletions from, additions to, and/orsubstitutions of one or more amino acid of an anti-B7RP1 antibody.

The term “polypeptide fragment” refers to a polypeptide that has anamino-terminal deletion, a carboxyl-terminal deletion, and/or aninternal deletion. In certain embodiments, fragments are at least 5 toabout 500 amino acids long. It will be appreciated that in certainembodiments, fragments are at least 5, 6, 8, 10, 14, 20, 50, 70, 100,110, 150, 200, 250, 300, 350, 400, or 450 amino acids long. Particularlyuseful polypeptide fragments include functional domains, includingbinding domains particularly antigen-binding domains, especially whereinthe antigen is an epitope of human B7RP1. In the case of an anti-B7RP1antibody, useful fragments include but are not limited to a CDR region,a variable domain of a heavy or light chain, a portion of an antibodychain or just its variable region including two CDRs, and the like.

The term “specific binding agent” refers to a naturally occurring ornon-naturally occurring molecule that specifically binds to a target.Examples of specific binding agents include, but are not limited to,proteins, peptides, nucleic acids, carbohydrates, and lipids. In certainembodiments, a specific binding agent is an antibody.

The term “specific binding agent to B7RP1” refers to a specific bindingagent that specifically binds any portion of B7RP1. In certainembodiments, a specific binding agent to B7RP1 is an antibody that bindsspecifically to B7RP1.

By way of example, an antibody “binds specifically” to a target if theantibody, when labeled, can be competed away from its target by thecorresponding non-labeled antibody.

The term “immunologically functional immunoglobulin fragment” as usedherein refers to a polypeptide fragment that contains at least the CDRsof the immunoglobulin heavy and light chains. An immunologicallyfunctional immunoglobulin fragment of the invention is capable ofbinding to an antigen. In certain embodiments, the antigen is a ligandthat specifically binds to a receptor. In these embodiments, binding ofan immunologically functional immunoglobulin fragment of the inventionprevents binding of the ligand to its receptor, interrupting thebiological response resulting from ligand binding to the receptor. Inone embodiment, an immunologically functional immunoglobulin fragment ofthe invention binds specifically to B7RP1. Preferably, the fragmentbinds specifically to human B7RP1.

The term “naturally-occurring” or “native” as used herein and applied toan object refers to the fact that the object can be found in nature. Forexample, a polypeptide or polynucleotide sequence that is present in anorganism (including viruses) that can be isolated from a source innature and that has not been intentionally modified by man isnaturally-occurring. The term “non-naturally occurring” or “non-native”as used herein refers to a material that is not found in nature or thathas been structurally modified or synthesized by man. For example,“non-naturally occurring” can refer to a variant, such as apolynucleotide variant that can be produced using art-known mutagenesistechniques, or a polypeptide variant produced by such a polynucleotidevariant. Such variants include, for example, those produced bynucleotide substitutions, deletions or additions that may involve one ormore nucleotides. Polynucleotide variants can be altered in coding ornon-coding regions or both. Alterations in the coding regions mayproduce conservative or non-conservative amino acid substitutions,deletions, or additions Especially certain among these are silentsubstitutions, additions, deletions, and conservative substitutions,which do not alter the properties and activities of a B7RP1 antibody ofthe invention. One of skill in the art can readily determine how togenerate such a variant using methods well known in the art.

The term “operably linked” means that the components to which the termis applied are in a relationship that allows them to carry out theirinherent functions under suitable conditions. For example, a controlsequence “operably linked” to a protein coding sequence is ligatedthereto so that expression of the protein coding sequence is achievedunder conditions compatible with the transcriptional activity of thecontrol sequences.

The term “control sequence” as used herein refers to polynucleotidesequences that can effect expression, processing or intracellularlocalization of coding sequences to which they are operably linked. Thenature of such control sequences may depend upon the host organism. Inparticular embodiments, control sequences for prokaryotes may include apromoter, ribosomal binding site, and transcription terminationsequence. In other particular embodiments, control sequences foreukaryotes may include promoters comprising one or a plurality ofrecognition sites for transcription factors, transcription enhancersequences, transcription termination sequences and polyadenylationsequences. In certain embodiments, “control sequences” can includeleader sequences and/or fusion partner sequences.

The term “vector” includes a nucleic acid molecule capable of carryinginto a cell another nucleic acid to which it has been linked. One typeof vector is a “plasmid”, which refers to a circular double stranded DNAloop into which additional DNA segments may be ligated. Another type ofvector is a viral vector, wherein additional DNA segments may be ligatedinto the viral genome. Certain vectors are capable of autonomousreplication in a host cell into which they are introduced (e.g.,bacterial vectors having a bacterial origin of replication and episomalmammalian vectors). Other vectors (e.g., non-episomal mammalian vectors)can be integrated into the genome of a host cell upon introduction intothe host cell and thereby are replicated along with the host genome.Moreover, certain vectors are capable of directing the expression ofgenes to which they are operatively linked. Such vectors are referred toherein as “recombinant expression vectors” (or simply, “expressionvectors”). In general, expression vectors useful in the practice ofrecombinant DNA techniques are often in the form of plasmids. In thepresent specification, “plasmid” and “vector” may be usedinterchangeably as the plasmid is the most commonly used form of vector.However, the invention is intended to include such other forms ofexpression vectors, such as viral vectors (e.g., replication defectiveretroviruses, adenoviruses and adeno-associated viruses), which serveequivalent functions.

The phrase “recombinant host cell” (or simply “host cell”) includes acell into which a recombinant expression vector has been introduced. Itwill be understood by those of skill in the art that such terms areintended to refer not only to the particular subject cell but to theprogeny of such a cell. Because certain modifications may occur insucceeding generations due to either mutation or environmentalinfluences, such progeny may not, in fact, be identical to the parentcell, but are still included within the scope of the term “host cell” asused herein. A wide variety of host expression systems can be used toexpress the antibodies of the present invention including bacterial,yeast, baculoviral and mammalian expression systems (as well as phagedisplay expression systems). An example of a suitable bacterialexpression vector is pUC19. To express an antibody recombinantly, a hostcell is transfected with one or more recombinant expression vectorscarrying DNA fragments encoding the immunoglobulin light and heavychains of the antibody such that the light and heavy chains areexpressed in the host cell and can be secreted into the medium in whichthe host cells are cultured, from which medium the antibodies can berecovered. Standard recombinant DNA methodologies are used to obtainantibody heavy and light chain genes, incorporate these genes intorecombinant expression vectors and introduce the vectors into hostcells, such as those described in Sambrook et al., 2001, MOLECULARCLONING, A LABORATORY MANUAL, Cold Spring Harbor Laboratories, Ausubel,F. M. et al. (eds.), CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, GreenePublishing Associates, (1989) and in U.S. Pat. No. 4,816,397 to Boss etal.

The term “transduction” is used to refer to the transfer of genes fromone bacterium to another, usually by a phage. “Transduction” also refersto the acquisition and transfer of eukaryotic cellular sequences byretroviruses.

The term “transfection” is used to refer to the uptake of foreign orexogenous DNA by a cell, and a cell has been “transfected” when theexogenous DNA has been introduced inside the cell membrane. A number oftransfection techniques are well known in the art and are disclosedherein. See, e.g., Graham et al., 1973, Virology 52: 456; Sambrook etal., 2001, MOLECULAR CLONING, A LABORATORY MANUAL, Cold Spring HarborLaboratories; Davis et al., 1986, BASIC METHODS IN MOLECULAR BIOLOGY,Elsevier; and Chu et al., 1981, Gene 13: 197. Such techniques can beused to introduce one or more exogenous DNA moieties into suitable hostcells.

The term “transformation” as used herein refers to a change in a cell'sgenetic characteristics, and a cell has been transformed when it hasbeen modified to contain a new DNA. For example, a cell is transformedwhere it is genetically modified from its native state. Followingtransfection or transduction, the transforming DNA may recombine withDNA from the cell by physically integrating into a chromosome of thecell, or may be maintained transiently as an episomal element withoutbeing replicated, or may replicate independently as a plasmid A cell isconsidered to have been stably transformed when the DNA is replicatedwith the division of the cell.

The term “antigen” refers to a molecule or a portion of a moleculecapable of being bound by a selective binding agent, such as anantibody, and additionally capable of being used in an animal to produceantibodies capable of binding to an epitope of that antigen. An antigenmay have one or more epitopes.

In certain embodiments, antibody variants include glycosylation variantswherein the number and/or type of glycosylation site has been alteredcompared to the amino acid sequences of the parent polypeptide. Incertain embodiments, protein variants comprise a greater or a lessernumber of N-linked glycosylation sites than the native protein. AnN-linked glycosylation site is characterized by the sequence:Asn-Xaa-Ser or Asn-Xaa-Thr, wherein the amino acid residue designated asXaa may be any amino acid residue except proline. The substitution ofamino acid residues to create this sequence provides a potential newsite for the addition of an N-linked carbohydrate chain. Alternatively,substitutions that eliminate this sequence will remove an existingN-linked carbohydrate chain. Also provided is a rearrangement ofN-linked carbohydrate chains wherein one or more N-linked glycosylationsites (typically those that are naturally occurring) are eliminated andone or more new N-linked sites are created. Additional antibody variantsinclude cysteine variants wherein one or more cysteine residues aredeleted from or substituted for another amino acid (e.g., serine)compared to the parent amino acid sequence. Cysteine variants may beuseful when antibodies must be refolded into a biologically activeconformation such as after the isolation of insoluble inclusion bodies.Cysteine variants generally have fewer cysteine residues than the nativeprotein, and typically have an even number to minimize interactionsresulting from unpaired cysteines.

In additional embodiments, antibody variants can include antibodiescomprising a modified Fc fragment or a modified heavy chain constantregion. An Fc fragment, which stands for “fragment that crystallizes,”or a heavy chain constant region can be modified by mutation to conferon an antibody altered binding characteristics. See, for example, Burtonand Woof, 1992, Advances in Immunology 51: 1-84; Ravetch and Bolland,2001, Annu. Rev. Immunol. 19: 275-90; Shields et al., 2001, Journal ofBiol. Chem. 276: 6591-6604; Telleman and Junghans, 2000, Immunology 100:245-251; Medesan et al., 1998, Eur. J. Immunol. 28: 2092-2100; all ofwhich are incorporated herein by reference). Such mutations can includesubstitutions, additions, deletions, or any combination thereof, and aretypically produced by site-directed mutagenesis using one or moremutagenic oligonucleotide(s) according to methods described herein, aswell as according to methods known in the art (see, for example,Sambrook et al., MOLECULAR CLONING: A LABORATORY MANUAL, 3rd Ed., 2001,Cold Spring Harbor, N.Y. and Berger and Kimmel, METHODS IN ENZYMOLOGY,Volume 152, Guide to Molecular Cloning Techniques, 1987, Academic Press,Inc., San Diego, Calif., which are incorporated herein by reference).

According to certain embodiments, amino acid substitutions may (1)reduce susceptibility to proteolysis, (2) reduce susceptibility tooxidation, (3) alter binding affinity, and/or (4) confer or modify otherphysicochemical or functional properties on such polypeptides. Accordingto certain embodiments, single or multiple amino acid substitutions (incertain embodiments, conservative amino acid substitutions) may be madein the naturally occurring sequence (in certain embodiments, in theportion of the polypeptide outside the domain(s) forming intermolecularcontacts). In certain embodiments, a conservative amino acidsubstitution typically does not substantially change the structuralcharacteristics of the parent sequence (e.g., a replacement amino acidshould disrupt or tend to disrupt secondary structure that characterizesa parent sequence, such as a helix). Examples of art-recognizedpolypeptide secondary and tertiary structures are described in PROTEINS,STRUCTURES AND MOLECULAR PRINCIPLES, (Creighton, Ed.), 1984, W. H.Freeman and Company, New York; INTRODUCTION TO PROTEIN STRUCTURE (C.Branden and J. Tooze, eds.), 1991, Garland Publishing, New York, N.Y.;and Thornton et al., 1991, Nature 354:105, each of which areincorporated herein by reference.

“Antibody” or “antibody peptide(s)” refer to an intact antibody, or abinding fragment thereof that competes with the intact antibody forspecific binding. In certain embodiments, binding fragments are producedby recombinant DNA techniques. In additional embodiments, bindingfragments are produced by enzymatic or chemical cleavage of intactantibodies. Binding fragments include, but are not limited to, F(ab),F(ab′), F(ab′)₂, Fv, and single-chain antibodies.

The invention provides antibodies that comprise a heavy chain and alight chain, wherein the heavy and light chains together form an antigenbinding structure capable of specifically binding B7RP1. A full-lengthheavy chain includes a variable region domain, V_(H), and three constantregion domains, C_(H)1, C_(H)2, and C_(H)3. The V_(H) domain is at theamino-terminus of the polypeptide, and the C_(H)3 domain is at thecarboxyl-terminus. The term “heavy chain”, as used herein, encompasses afull-length heavy chain and fragments thereof. A full-length light chainincludes a variable region domain, V_(L), and a constant region domain,C_(L). Like the heavy chain, the variable region domain of the lightchain is at the amino-terminus of the polypeptide The term “lightchain”, as used herein, encompasses a full-length light chain andfragments thereof. A F(ab) fragment is comprised of one light chain andthe C_(H)1 and variable regions of one heavy chain. The heavy chain of aF(ab) molecule cannot form a disulfide bond with another heavy chainmolecule. A F(ab′) fragment contains one light chain and one heavy chainthat contains more of the constant region, between the C_(H)1 and C_(H)2domains, such that an interchain disulfide bond can be formed betweentwo heavy chains to form a F(ab′)₂ molecule. The Fv region comprises thevariable regions from both the heavy and light chains, but lacks theconstant regions. Single-chain antibodies are Fv molecules in which theheavy and light chain variable regions have been connected by a flexiblelinker to form a single polypeptide chain, which forms anantigen-binding region. Single chain antibodies are discussed in detailin International Patent Application Publication No. WO 88/01649 and U.S.Pat. Nos. 4,946,778 and 5,260,203.

A bivalent antibody other than a “multispecific” or “multifunctional”antibody, in certain embodiments, is understood to comprise bindingsites having identical antigenic specificity.

In assessing antibody binding and specificity according to theinvention, an antibody substantially inhibits adhesion of a ligand to areceptor when an excess of antibody reduces the quantity of ligand boundto receptor by at least about 20%, 40%, 60%, 80%, 85%, or more (asmeasured, inter alia, using an in vitro competitive binding assay).

By “neutralizing antibody” is meant an antibody molecule that is able toblock or substantially reduce an effector function of a target antigento which it binds. Accordingly, a “neutralizing” anti-B7RP1 antibody iscapable of blocking or substantially reducing an effector function, suchas receptor binding and/or elicitation of a cellular response, of B7RP1.“Substantially reduce” is intended to mean at least about 60%, at leastabout 70%, at least about 75%, at least about 80%, at least about 85%,or at least about 90% reduction of an effector function of the targetantigen (e.g., human B7RP1).

The term “epitope” includes any site on an antigen that is capable ofspecific binding to an immunoglobulin or T-cell receptor. In certainembodiments, epitope determinants include chemically active surfacegroupings of molecules such as amino acids, sugar side chains,phosphoryl groups, or sulfonyl groups, and, in certain embodiments, mayhave specific three-dimensional structural characteristics, and/orspecific charge characteristics. An epitope is a region of an antigenthat is bound by an antibody. In certain embodiments, an antibody issaid to specifically bind an antigen when it preferentially recognizesits target antigen in a complex mixture of proteins and/ormacromolecules. In certain embodiments, an antibody is said tospecifically bind an antigen when the equilibrium dissociation constantis about 10⁻⁶ M, 10⁻⁷ M, 10⁻⁸ M, 10⁻⁹ M, 10⁻¹⁰ M, 10⁻¹¹ M, 10⁻¹² M, orless than about 10⁻¹²M.

An antibody binds “essentially the same epitope” as a referenceantibody, when the two antibodies recognize identical or stericallyoverlapping epitopes. The most widely used and rapid methods fordetermining whether two antibodies bind to identical or stericallyoverlapping epitopes are competition assays, which can be configured inall number of different formats, using either labeled antigen or labeledantibody. Usually, the antigen is immobilized on a substrate, and theability of unlabeled antibodies to block the binding of labeledantibodies is measured using radioactive isotopes or enzyme labels.

The term “agent” is used herein to denote a chemical compound, a mixtureof chemical compounds, a biological macromolecule, or an extract madefrom biological materials.

As used herein, the terms “label” or “labeled” refers to incorporationof a detectable marker, e.g., by incorporation of a radiolabeled aminoacid or attachment to a polypeptide of biotin moieties that can bedetected by labeled avidin (e.g., streptavidin comprising a detectablemarker such as a fluorescent marker, a chemiluminescent marker or anenzymatic activity that can be detected by optical or colorimetricmethods). In certain embodiments, the label can also be therapeutic.Various methods of labeling polypeptides and glycoproteins are known inthe art and may be used advantageously in the methods disclosed herein.Examples of labels for polypeptides include, but are not limited toradioisotopes or radionuclides such as ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, ^(99m)Tc,¹¹¹In, ¹²⁵I, and ¹³³I, fluorescent labels (e.g., fluoresceinisothiocyanate or FITC, rhodamine, or lanthanide phosphors), enzymaticlabels (e.g., horseradish peroxidase, β-galactosidase, luciferase,alkaline phosphatase), chemiluminescent labels, hapten labels such asbiotinyl groups, and predetermined polypeptide epitopes recognized by asecondary reporter (e.g., leucine zipper pair sequences, binding sitesfor secondary antibodies, metal binding domains, or epitope tags). Incertain embodiments, labels are attached by spacer arms (such as(CH₂)_(n), where n<about 20) of various lengths to reduce potentialsteric hindrance.

The term “biological sample”, as used herein, includes, but is notlimited to, any quantity of a substance from a living thing or formerlyliving thing. Such living things include, but are not limited to,humans, mice, monkeys, rats, rabbits, and other animals. Such substancesinclude, but are not limited to, blood, serum, urine, cells, organs,tissues, bone, bone marrow, lymph nodes, and skin.

The term “pharmaceutical agent or drug” as used herein refers to achemical compound or composition capable of inducing a desiredtherapeutic effect when properly administered to a patient. Theexpression “pharmaceutically effective amount” in reference to apharmaceutical composition comprising one or a plurality of theantibodies of the invention is understood to mean, according to theinvention, an amount of the said pharmaceutical composition that iscapable of abolishing, in a patient, the decrease in the sensitivitythreshold to external stimuli with a return of this sensitivitythreshold to a level comparable to that observed in healthy subjects.

A “disorder” is any condition that would benefit from treatmentaccording to the present invention. “Disorder” and “condition” are usedinterchangeably herein and include chronic and acute immune systemdisorders or immune system diseases associated with inappropriate immuneresponse, including those pathological conditions which predispose themammal to the disorder in question. A number of conditions and disordersthat would benefit from the treatment according to the present inventionare described, for example, in International Patent Application NoPCT/US00/01871 (Publication No. WO 00/46240), the disclosure of which isincorporated by reference in its entirety.

The terms “immune system disease” and “immune system condition”encompass any medical condition or disorder associated with increasedlevels of B7RP1, increased sensitivity to B7RP1, or T-cell mediateddiseases, including, but not limited to, autoimmune disease, graftsurvival, bone marrow and organ transplantation, allosensitization dueto blood transfusions, toxic shock syndrome, T-cell dependent B-cellmediated diseases, chronic inflammatory diseases associated with chronicimmune cell dysfunction, lymphoproliferative disorders (such as multiplemyeloma, Waldenstom's macroglobulinemia, and crioglobulinemias), andcancer. Non-limiting examples of autoimmune diseases include systemiclupus erythematosis, rheumatoid arthritis, immune thrombocytopenicpurpura (ITP), multiple sclerosis, diabetes, and psoriasis. Non-limitingexamples of chronic inflammatory diseases include inflammatory boweldisease (such as Crohn's disease and ulcerative colitis), Grave'sdisease, Hashimoto's thyroiditis, and diabetes mellitus.

The terms “immune system disease” and “immune system condition” alsoencompass any clinical condition that would be ameliorated by theinhibition of antibody production, such as hypersensitivity reactions.Hypersensitivity reactions can be caused, for example, by hay fever,allergies, asthma, atopy, and acute edema. Non-limiting examples ofdiseases that cause antibody-mediated hypersensitivity reactions includesystemic lupus erythematosis, arthritis (such as rheumatoid arthritis,reactive arthritis, psoriatic arthritis), nephropathies (such asglomerulo-nephritis, membranous, mesangiocapillary, focal segmental,focal necrotizing, crescentic, and proliferative nephropathies such astubulopathies), skin disorders (such as pemphigus and pemphigoid,erythema nodosum), endocrinopathies (such as thyroiditis, Grave'sdisease, Hashimoto's disease, insulin dependent diabetes mellitus),various pneumopathies (such as extrinsic alveolitis), variousvasculopathies, coeliac disease, diseases with aberrant production ofIgA, many anemias and thrombocytopenias, Guillain-Barre Syndrome, andmyasthenia gravis.

As used herein, the terms “effective amount” and “therapeuticallyeffective amount” when used with reference to a vehicle- or apharmaceutical composition comprising one or more anti-human B7RP1 humanantibodies refers to an amount or dosage sufficient to produce a desiredresult (i.e., where for therapy with the vehicle- or anti-human B7RP1human antibodies of the present invention the desired result is thedesired modulation of T-cell responses, for example) or to support anobservable decrease in the level of one or more biological activities ofB7RP1. More specifically, a therapeutically effective amount is anamount of the anti-human B7RP1 human antibody(ies) sufficient toinhibit, for some period of time, one or more of the clinically definedpathological processes associated with the condition at issue, e.g.immune disorders and diseases, in a subject treated in vivo with theagent. In the present invention, an “effective amount” of an anti-B7RP1antibody may modulate T-cell responses in a patient. In the methods ofthe present invention, the term “control” and grammatical variantsthereof, are used to refer to the prevention, partial or completeinhibition, reduction, delay or slowing down of an unwanted event, e.g.immune response. The effective amount may vary depending on the specificvehicle- or anti-human B7RP1 human antibody(ies) selected, and is alsodependent on a variety of factors and conditions related to the subjectto be treated and the severity of the disorder. For example, if thevehicle- or anti-human B7RP1 human antibody(ies) is to be administeredin vivo, factors such as the age, weight and health of the patient aswell as dose response curves and toxicity data obtained in preclinicalanimal work would be among those considered. If the agent is to becontacted with the cells in vitro, one would also design a variety ofpre-clinical in vitro studies to assess such parameters as uptake,half-life, dose, toxicity, etc. The determination of an effective amountor a therapeutically effective amount for a given agent is well withinthe ability of those skilled in the art.

As used herein, the terms “B7 related protein-1” and “B7RP1” are definedas all mammalian species of native sequence B7RP1, which is described inInternational Patent Application Publication No. WO 00/46240, which isincorporated herein by reference.

As used herein, “substantially pure” or “substantially purified” means acompound or species that is the predominant species present (i.e., on amolar basis it is more abundant than any other individual species in thecomposition). In certain embodiments, a substantially purified fractionis a composition wherein the species comprises at least about 50 percent(on a molar basis) of all macromolecular species present. In certainembodiments, a substantially pure composition will comprise more thanabout 80%, 85%, 90%, 95%, or 99% of all macromolar species present inthe composition. In certain embodiments, the species is purified toessential homogeneity (contaminant species cannot be detected in thecomposition by conventional detection methods) wherein the compositionconsists essentially of a single macromolecular species.

The term “patient” includes human and animal subjects.

“Treatment” or “treat” refers to both therapeutic treatment andprophylactic or preventative measures. Those in need of treatmentinclude those already with the disorder as well as those prone to havethe disorder or those in which the disorder is to be prevented.

Unless otherwise required by context, singular terms shall includepluralities and plural terms shall include the singular.

According to certain embodiments of the invention, antibodies directedto B7RP1 may be used to treat immune system disorders and immune systemdiseases, including but not limited to, those mentioned above.

In one aspect of the invention are provided fully human monoclonalantibodies raised against and having biological and immunologicalspecificity for binding to human B7RP1. In another aspect the inventionprovides nucleic acids comprising nucleotide sequences encoding aminoacid sequences for heavy and light chain immunoglobulin molecules,particularly sequences corresponding to the variable regions thereof.Particular embodiments of this aspect of the invention are sequencescorresponding to complementarity determining regions (CDRs),specifically from CDR1 through CDR3, of the heavy and light chainsprovided by the invention. In yet another aspect the invention provideshybridoma cells and cell lines that express the immunoglobulin moleculesand antibodies, such as monoclonal antibodies of the invention. Theinvention also provides biologically and immunologically purifiedpreparations of antibodies, such as monoclonal antibodies raised againstand having biological and immunological specificity for binding to humanB7RP1.

The ability to clone and reconstruct megabase-sized human loci in yeastartificial chromosomes (YACs) and to introduce them into the mousegermline provides an advantageous approach to elucidating the functionalcomponents of very large or crudely mapped loci as well as generatinguseful models of human disease. Furthermore, the utilization of suchtechnology for substitution of mouse loci with their human equivalentsprovides unique insights into the expression and regulation of humangene products during development, their communication with othersystems, and their involvement in disease induction and progression.

An important practical application of such a strategy is the“humanization” of the mouse humoral immune system. Introduction of humanimmunoglobulin (Ig) loci into mice in which the endogenous Ig genes havebeen inactivated offers the opportunity to study mechanisms underlyingprogrammed expression and assembly of antibodies as well as their rolein B-cell development. Furthermore, such a strategy provides a sourcefor production of fully human monoclonal antibodies (MAbs).

The term “human antibody” includes antibodies having variable andconstant regions substantially corresponding to human germlineimmunoglobulin sequences. In certain embodiments, human antibodies areproduced in non-human mammals, including, but not limited to, rodents,such as mice and rats, and lagomorphs, such as rabbits. In certainembodiments, human antibodies are produced in hybridoma cells In certainembodiments, human antibodies are produced recombinantly.

The term “recombinant” in reference to an antibody includes antibodiesthat are prepared, expressed, created or isolated by recombinant means.Representative examples include antibodies expressed using a recombinantexpression vector transfected into a host cell, antibodies isolated froma recombinant, combinatorial human antibody library, antibodies isolatedfrom an animal (e.g., a mouse) that is transgenic for humanimmunoglobulin genes (see e.g., Taylor, et al., 1992, Nucl. Acids Res.,20:6287-6295); or antibodies prepared, expressed, created or isolated byany means that involves splicing of human immunoglobulin gene sequencesto other DNA sequences. Such recombinant human antibodies have variableand constant regions derived from human germline immunoglobulinsequences.

Human antibodies have at least three advantages over non-human andchimeric antibodies for use in human therapy:

1) because the effector portion of the antibody is human, it mayinteract better with the other parts of the human immune system (e.g.,destroy the target cells more efficiently by complement-dependentcytotoxicity (CDC) or antibody-dependent cellular cytotoxicity (ADCC));

2) the human immune system should not recognize the human antibody asforeign, and, therefore the antibody response against such an injectedantibody should be less than against a totally foreign non-humanantibody or a partially foreign chimeric antibody;

3) injected non-human antibodies have been reported to have a half-lifein the human circulation much shorter than the half-life of humanantibodies. Injected human antibodies will have a half-life essentiallyidentical to naturally occurring human antibodies, allowing smaller andless frequent doses to be given.

Thus, fully human antibodies are expected to minimize the immunogenicand allergic responses intrinsic to mouse or mouse-derivatized MAbs, andto thereby increase the efficacy and safety of the administeredantibodies. Fully human antibodies of the invention, therefore, can beused in the treatment of diseases and disorders associated withinappropriate immune response, the treatment thereof requiring repeatedantibody administration. Thus, one particular advantage of theanti-B7RP1 antibodies of the invention is that the antibodies are fullyhuman and can be administered to patients in a non-acute manner whileminimizing adverse reactions commonly associated with human anti-mouseantibodies or other previously described non-fully human antibodies fromnon-human species.

One skilled in the art can engineer mouse strains deficient in mouseantibody production with large fragments of the human Ig loci so thatsuch mice produce human antibodies in the absence of mouse antibodies.Large human Ig fragments may preserve the large variable gene diversityas well as the proper regulation of antibody production and expression.By exploiting the mouse cellular machinery for antibody diversificationand selection and the lack of immunological tolerance to human proteins,the reproduced human antibody repertoire in these mouse strains yieldshigh affinity antibodies against any antigen of interest, includinghuman antigens. Using the hybridoma technology, antigen-specific humanMAbs with the desired specificity may be produced and selected.

Transgenic animals (e.g., mice) can also be used to produce humanantibodies in the absence of endogenous immunoglobulin production. Forexample, transfer of the human germ-line immunoglobulin gene array insuch germ-line mutant mice will result in the production of humanantibodies upon antigen challenge (see, e.g., Jakobovits et al., 1993,Proc. Natl. Acad. Sci. USA 90:2551-2555; Jakobovits et al., 1993, Nature362:255-258; Bruggemann et al., 1993, Year in Immun. 7:33, 1994, Nature148:1547-1553) and, 1996, Nature Biotechnology 14:826; Gross et al.,2000, Nature 404:995-999; and U.S. Pat. Nos. 5,877,397, 5,874,299,5,814,318, 5,789,650, 5,770,429, 5,661,016, 5,633,425, 5,625,126,5,569,825, and 5,545,806, (each of which is incorporated herein byreference in its entirety for all purposes)). Human antibodies can alsobe produced in phage display libraries (Hoogenboom and Winter, 1992, J.Mol. Biol 227:381; Marks et al., 1991, J. Mol. Biol. 222:581). Thetechniques of Cole et al. and Boerner et al. are also available for thepreparation of human monoclonal antibodies (Cole et al., 1985,MONOCLONAL ANTIBODIES AND CANCER THERAPY Alan R. Liss, p. 77; andBoerner et al., 1991, J. Immunol. 147:86-95).

Recombinant human antibodies may also be subjected to in vitromutagenesis (or, when an animal transgenic for human Ig sequences isused, in vivo somatic mutagenesis) and, thus, the amino acid sequencesof the V_(H) and V_(L) regions of the recombinant antibodies aresequences that, while derived from those related to human germline V_(H)and V_(L) sequences, may not naturally exist within the human antibodygermline repertoire in vivo.

In certain embodiments, the skilled artisan can use constant regionsfrom species other than human along with the human variable region(s) insuch mice to produce chimeric antibodies.

A bispecific or bifunctional antibody typically is an artificial hybridantibody having two different heavy chain/light chain pairs and twodifferent binding sites. Bispecific antibodies may be produced by avariety of methods including, but not limited to, fusion of hybridomasor linking of F(ab′) fragments, See, e.g., Songsivilai & Lachmann, 1990,Clin. Exp Immunol. 79: 315-321; Kostelny et al., 1992, J. Immunol.148:1547-1553.

The invention provides antibodies that bind to human B7RP1. Theseantibodies can be produced by immunization with full-length B7RP1 orfragments thereof. The antibodies of the invention can be polyclonal ormonoclonal, and/or may be recombinant antibodies. In preferredembodiments, antibodies of the invention are human antibodies prepared,for example, by immunization of transgenic animals capable of producinghuman antibodies (see, for example, International Patent Application,Publication WO 93/12227).

The complementarity determining regions (CDRs) of the light chain andheavy chain variable regions of anti-B7RP1 antibodies of the inventioncan be grafted to framework regions (FRs) from the same, or another,species. In certain embodiments, the CDRs of the light chain and heavychain variable regions of anti-B7RP1 antibody may be grafted toconsensus human FRs. To create consensus human FRs, FRs from severalhuman heavy chain or light chain amino acid sequences are aligned toidentify a consensus amino acid sequence. The FRs of the anti-B7RP1antibody heavy chain or light chain can be replaced with the FRs from adifferent heavy chain or light chain. Rare amino acids in the FRs of theheavy and light chains of anti-B7RP1 antibody typically are notreplaced, while the rest of the FR amino acids can be replaced. Rareamino acids are specific amino acids that are in positions in which theyare not usually found in FRs. The grafted variable regions fromanti-B7RP1 antibodies of the invention can be used with a constantregion that is different from the constant region of anti-B7RP1antibody. Alternatively, the grafted variable regions are part of asingle chain Fv antibody. CDR grafting is described, e.g., in U.S. Pat.Nos. 6,180,370, 5,693,762, 5,693,761, 5,585,089, and 5,530,101, whichare hereby incorporated by reference for any purpose.

Antibodies of the invention can be prepared using transgenic mice thathave a substantial portion of the human antibody producing locusinserted in antibody-producing cells of the mice, and that are furtherengineered to be deficient in producing endogenous, murine, antibodies.Such mice are capable of producing human immunoglobulin molecules andantibodies and do not produce or produce substantially reduced amountsof murine immunoglobulin molecules and antibodies. Technologies utilizedfor achieving this result are disclosed in the patents, applications,and references disclosed in the specification herein. In certainembodiments, the skilled worker may employ methods as disclosed inInternational Patent Application Publication No. WO 98/24893, which ishereby incorporated by reference for any purpose. See also Mendez etal., 1997, Nature Genetics 15:1.46-156, which is hereby incorporated byreference for any purpose.

The monoclonal antibodies (mAbs) of the invention can be produced by avariety of techniques, including conventional monoclonal antibodymethodology, e.g., the standard somatic cell hybridization technique ofKohler and Milstein (1975, Nature 256:495). Other techniques forproducing monoclonal antibodies may be employed, e.g., viral oroncogenic transformation of B-lymphocytes.

An exemplary animal system for preparing hybridomas is the mouse.Hybridoma production in the mouse is known in the art and immunizationprotocols and techniques for isolation of immunized splenocytes forfusion are also known in the art. Fusion partners (e.g., murine myelomacells) and fusion procedures are also known.

In a certain embodiment, human monoclonal antibodies directed againstB7RP1 can be generated using transgenic mice carrying parts of the humanimmune system rather than the mouse system. These transgenic mice,referred to herein as “HuMab” mice, contain a human immunoglobulin geneminilocus that encodes unrearranged human heavy (μ and γ) and κ lightchain immunoglobulin sequences, together with targeted mutations thatinactivate the endogenous μ and κ chain loci (Lonberg et al., 1994,Nature 368:856-859). Accordingly, the mice exhibit reduced expression ofmouse IgM or κ and in response to immunization, the introduced humanheavy chain and light chain transgenes undergo class switching andsomatic mutation to generate high affinity human IgG κ monoclonalantibodies (Lonberg et al., supra.; Lonberg and Huszar, 1995, Intern.Rev. Immunol. 13:65-93; Harding and Lonberg, 1995, Ann. N.Y. Acad. Sci.764:536-546). The preparation of HuMab mice is described in detail inTaylor et al., 1992, Nucleic Acids Res. 20:6287-6295; Chen et al., 1993,International Immunology 5:647-656; Tuaillon et at, 1994, J. Immunol.152:2912-2920; Lonberg et al., 1994, Nature 368:856-859; Lonberg, 1994,Handbook of Exp. Pharmacology 113:49-101; Taylor et at, 1994,International Immunology 6:579-591; Lonberg & Huszar, 1995, Intern. Rev.Immunol. 13:65-93; Harding & Lonberg, 1995, Ann. N.Y. Acad. Sci.764:536-546; Fishwild et al., 1996, Nature Biotechnology 14:845-851, thecontents of all of which are hereby incorporated by reference in theirentirety. See further U.S. Pat. Nos. 5,545,806; 5,569,825; 5,625,126;5,633,425; 5,789,650; 5,877,397; 5,661,016; 5,814,318; 5,874,299; and5,770,429; all to Lonberg and Kay, as well as U.S. Pat. No. 5,545,807 toSurani et al.; International Patent Application Publication Nos. WO93/1227, published Jun. 24, 1993; WO 92/22646, published Dec. 23, 1992;and WO 92/03918, published Mar. 19, 1992, the disclosures of all ofwhich are hereby incorporated by reference in their entirety.Alternatively, transgenic mice strains described in the Examples belowcan be used to generate human anti-B7RP1 antibodies.

The present invention provides human monoclonal antibodies that arespecific for and neutralize bioactive human B7RP1 polypeptides. Alsoprovided are antibody heavy and light chain amino acid sequences whichare highly specific for and neutralize B7RP1 polypeptides when they arebound to them. This high specificity enables the anti-human B7RP1 humanantibodies, and human monoclonal antibodies with like specificity, to beeffective immunotherapy for B7RP1 associated diseases.

In one aspect, the invention provides isolated human antibodies thatbind the same or essentially the same epitope as the 16H antibodyprovided herein.

In one aspect, the invention provides isolated human antibodiescomprising at least one of the amino acid sequences shown in SEQ ID NOS:1-40 or 44-58 that binds a B7RP1 polypeptide epitope with high affinityand has the capacity to antagonize B7RP1 polypeptide activity. Theseantibodies may bind the same or essentially the same epitope as theanti-B7RP1 antibodies shown in the Examples herein.

In certain embodiments, the isolated antibodies bind to B7RP1polypeptide with a dissociation constant (K_(D)) of about 10⁻⁶ M, 10⁻⁷M, 10⁴ M, 10⁻⁹ M, 10⁻¹⁰ M, 10⁻¹¹ M or less and inhibits B7RP1 inducedsurvival in an in vitro neutralization assay with an EC₅₀ of about 10⁻⁶M, 10⁻⁷ M, 10⁻⁸ M, 10⁻⁹ M or less Examples of anti-human B7RP1 humanantibodies that meet the aforementioned binding and neutralizationcriteria are provided herein.

In certain embodiments, anti-human B7RP1 human antibodies of theinvention are referred to herein as 16H, 16Hg (germline), 5D, 2H, 2Hg(germline), 15H, 41H, and 43H. Antibody 16H comprises V_(L) and V_(H)polypeptide sequences as shown in SEQ ID NO: 7 and SEQ ID NO: 1,respectively. Antibody 16Hg comprises a variable light chain (V_(L)) andvariable heavy chain (V_(H)) polypeptide sequences as shown in SEQ IDNO: 1 and SEQ ID NO: 8, respectively. Antibody 5D comprises V_(L) andV_(H) polypeptide sequences as shown in SEQ ID NO: 2 and SEQ ID NO: 9,respectively. Antibody 2H comprises V_(L) and V_(H) polypeptidesequences as shown in SEQ ID NO: 3 and SEQ ID NO: 10, respectively.Antibody 2Hg comprises V_(L) and V_(H) polypeptide sequences as shown inSEQ ID NO: 3 and SEQ ID NO: 11, respectively. Antibody 15H comprises V₁and V_(H) polypeptide sequences as shown in SEQ ID NO: 4 and SEQ ID NO:12, respectively. Antibody 41H comprises V_(L) and V_(H) polypeptidesequences as shown in SEQ ID NO: 5 and SEQ ID NO: 13, respectively.Antibody 43H comprises V_(L) and V_(H) polypeptide sequences as shown inSEQ ID NO: 6 and SEQ ID NO: 14, respectively. The properties of theanti-human B7RP1 human antibodies of the present invention arespecifically disclosed in the Examples.

Particularly notable is the high affinity for B7RP1 polypeptide and highcapacity to antagonize B7RP1 polypeptide activity demonstrated herein.

The dissociation constant (K_(D)) of an anti-human B7RP1 human antibodycan be determined by surface plasmon resonance as generally described inthe Examples below. Generally, surface plasmon resonance analysismeasures real-time binding interactions between ligand (recombinantB7RP1 polypeptide immobilized on a biosensor matrix) and analyte(antibodies in solution) by surface plasmon resonance (SPR) using theBIAcore® system (Pharmacia Biosensor, Piscataway, N.J.). Surface plasmonanalysis can also be performed by immobilizing the analyte (antibodieson a biosensor matrix) and presenting the ligand (recombinant V insolution). The dissociation constant (K_(D)) of an anti-human B7RP1human antibody can also be determined by using KinExA methodology. Incertain embodiments of the invention, the antibodies bind to B7RP1 witha K_(D) of approximately 10⁻⁵ M, 10⁻⁶ M, 10⁻⁷ M, 10⁻⁸ M, 10⁻⁹ M, 10⁻¹⁰M, 10⁻¹¹ M, or 10⁻¹² M. The term “K_(D)”, as used herein, is intended torefer to the dissociation constant of a particular antibody-antigeninteraction. For purposes of the present invention K_(D) was determinedas shown in the Examples below.

In certain embodiments, the antibodies of the invention are of the IgG1,IgG2, IgG3, or IgG4 isotype. The antibodies may be of the IgG2 or IgG1isotype. In other embodiments, the antibodies of the invention may be ofthe IgM, IgA, IgE, or IgD isotype. In certain embodiments of theinvention, the antibodies comprise a human kappa light chain and a humanIgG1, IgG2, IgG3, or IgG4 heavy chain. Expression of antibodies of theinvention comprising an IgG1 or an IgG2 heavy chain constant region isdescribed in the Examples below. In particular embodiments, the variableregions of the antibodies are ligated to a constant region other thanthe constant region for the IgG1, IgG2, IgG3, or IgG4 isotype. Incertain embodiments, the antibodies of the invention have been clonedfor expression in mammalian cells.

In certain embodiments, conservative modifications to the heavy chainsand light chains of anti-B7RP1 antibodies (and correspondingmodifications to the encoding nucleotides) will produce anti-B7RP1antibodies having functional and chemical characteristics similar tothose of the anti-B7RP1 antibodies disclosed herein. In contrast,substantial modifications in the functional and/or chemicalcharacteristics of anti-B7RP1 antibodies may be accomplished byselecting substitutions in the amino acid sequence of the heavy andlight chains that differ significantly in their effect on maintaining(a) the structure of the molecular backbone in the area of thesubstitution, for example, as a sheet or helical conformation, (b) thecharge or hydrophobicity of the molecule at the target site, or (c) thebulk of the side chain.

For example, a “conservative amino acid substitution” may involve asubstitution of a native amino acid residue with a normative residuesuch that there is little or no effect on the polarity or charge of theamino acid residue at that position. Furthermore, any native residue inthe polypeptide may also be substituted with alanine, as has beenpreviously described for “alanine scanning mutagenesis.”

Amino acid substitutions (whether conservative or non-conservative) canbe determined by those skilled in the art at the time such substitutionsare desired. In certain embodiments, amino acid substitutions can beused to identify those amino acid residues of an anti-B7RP1 antibodythat are involved in binding specificity and/or affinity of the antibodyfor B7RP1 (e.g. residues that are involved in binding of the antibody toa particular epitope), such as amino acid residues in CDR1, CDR2, and/orCDR3 regions of the light or heavy chains as described herein. Suchamino acid substitutions may increase or decrease the affinity of theanti-B7RP1 antibodies described herein.

Minor changes in an amino acid sequence such as deletion, addition orsubstitution of one, a few or even several amino acids may lead to anallelic form of the original protein which has substantially identicalproperties. Therefore, in addition to the antibodies specificallydescribed herein, other “substantially homologous” antibodies can bereadily designed and manufactured utilizing various recombinant DNAtechniques well known to those skilled in the art. In general,modifications of the genes may be readily accomplished by a variety ofwell-known techniques, such as site-directed mutagenesis. Therefore, thepresent invention contemplates “variant” or “mutant” anti-B7RP1 humanantibodies having substantially similar characteristics to theanti-B7RP1 human antibodies disclosed herein (See, for example, WO00/56772, all of which is hereby incorporated herein by reference).Thus, by the term “variant” or “mutant” in reference to an anti-B7RP1human antibody is meant any binding molecule (molecule X) (i) in whichthe hypervariable regions CDR1, CDR2, and CDR3 of the heavy chain or thehypervariable regions CDR1, CDR2, and CDR3 of the light chain taken as awhole are at least about 80% homologous, at least about 90% homologous,or at least about 95% homologous to the hypervariable regions as shownin SEQ ID NO: 15 through SEQ ID NO. 26 or SEQ ID NO: 27 through SEQ IDNO: 40, respectively, and (ii) wherein the variant or mutant is capableof inhibiting the activity of human B7RP1 to the same extent as areference anti-B7RP1 human antibody having framework regions identicalto those of molecule X. Such antibodies may bind to human B7RP1 or tomouse B7RP1 or both. The mouse B7RP1 sequence is described in WO00/46240, which is incorporated by reference.

Ordinarily, an anti-B7RP1 human antibody variant will have light and/orheavy chain CDRs, when taken as a whole, that are at least about 80%amino acid sequence identity, at least about 85% sequence identity, atleast about 90% sequence identity, at least about 91% sequence identity,at least about 92% sequence identity, at least about 9.3% sequenceidentity, at least about 94% sequence identity, at least about 95%sequence identity, at least about 96% sequence identity, at least about97% sequence identity, at least about 98% sequence identity, or at leastabout 99% amino acid sequence identity to the amino acid sequence asshown in SEQ ID NOS: 15 through SEQ ID NO. 26 and/or SEQ ID NOS: 27through SEQ ID NO. 40, respectively. Such antibodies may bind to humanB7RP1 or to mouse B7RP1 or to both.

An anti-B7RP1 human antibody variant will have a light chain variableregion, when taken as a whole, that has at least about 80% amino acidsequence identity, at least about 81% sequence identity, at least about82% sequence identity, at least about 83% sequence identity, at leastabout 84% sequence identity, at least about 85% sequence identity, atleast about 86% sequence identity, at least about 87% sequence identity,at least about 88% sequence identity, at least about 89% sequenceidentity, at least about 90% sequence identity, at least about 91%sequence identity, at least about 92% sequence identity, at least about93% sequence identity, at least about 94% sequence identity, at leastabout 95% sequence identity, at least about 96% sequence identity, atleast about 97% sequence identity, at least about 98% sequence identity,at least about 99% amino acid sequence identity to the amino acidsequence as shown in SEQ ID NOS: 1 through SEQ ID NO. 6, and/or a heavychain variable region, when taken as a whole, that has at least about70% amino acid sequence identity, at least about 75% sequence identity,at least about 80% sequence identity, at least about 81% sequenceidentity, at least about 82% sequence identity, at least about 83%sequence identity, at least about 84% sequence identity, at least about85% sequence identity, at least about 86% sequence identity, at leastabout 87% sequence identity, at least about 88% sequence identity, atleast about 89% sequence identity, at least about 90% sequence identity,at least about 91% sequence identity, at least about 92% sequenceidentity, at least about 93% sequence identity, at least about 94%sequence identity, at least about 95% sequence identity, at least about96% sequence identity, at least about 97% sequence identity, at leastabout 98% sequence identity, or at least about 99% amino acid sequenceidentity to the amino acid sequence as shown in SEQ ID NOS: 7 throughSEQ ID NO. 14. Such antibodies may bind to human B7RP1 and/or mouseB7RP1.

As will be appreciated by those of skill in the art, many of thepotential CDR-contact residues are amenable to substitution by otheramino acids and still allow the antibody to retain substantial affinityfor the antigen. Likewise, many of the framework residues not in contactwith the CDRs in the heavy and light chains can accommodatesubstitutions of amino acids from the corresponding positions from otherhuman antibodies, by human consensus amino acids, or from other mouseantibodies, without significant loss of the affinity ornon-immunogenicity of the human antibody. Selection of variousalternative amino acids may be used to produce versions of the disclosedanti-B7RP1 antibodies and fragments thereof that have varyingcombinations of affinity, specificity, non-immunogenicity, ease ofmanufacture, and other desirable properties.

A “variant” in reference to a polynucleotide is intended to refer to anucleic acid molecule having at least about 75% nucleic acid sequenceidentity with a polynucleotide sequence of the present invention.Ordinarily, a polynucleotide variant will have at least about 75%nucleic acid sequence identity, at least about 80% nucleic acid sequenceidentity, at least about 81% nucleic acid sequence identity, at leastabout 82% nucleic acid sequence identity, at least about 83% nucleicacid sequence identity, at least about 84% nucleic acid sequenceidentity, at least about 85% nucleic acid sequence identity, at leastabout 86% nucleic acid sequence identity, at least about 87% nucleicacid sequence identity, at least about 88% nucleic acid sequenceidentity, at least about 89% nucleic acid sequence identity, at leastabout 90% nucleic acid sequence identity, at least about 91% nucleicacid sequence identity, at least about 92% nucleic acid sequenceidentity, at least about 93% nucleic acid sequence identity, at leastabout 94% nucleic acid sequence identity, at least about 95% nucleicacid sequence identity, at least about 96% nucleic acid sequenceidentity, at least about 97% nucleic acid sequence identity, at leastabout 98% nucleic acid sequence identity, or at least about 99% nucleicacid sequence identity with a novel nucleic acid sequence disclosedherein.

In alternative embodiments, antibodies of the invention can be expressedin cell lines other than hybridoma cell lines. In these embodiments,sequences encoding particular antibodies can be used for transformationof a suitable mammalian host cell. According to these embodiments,transformation can be achieved using any known method for introducingpolynucleotides into a host cell, including, for example packaging thepolynucleotide in a virus (or into a viral vector) and transducing ahost cell with the virus (or vector) or by transfection procedures knownin the art, as exemplified by U.S. Pat. Nos. 4,399,216, 4,912,040,4,740,461, and 4,959,455 (all of which are hereby incorporated herein byreference for any purpose). Generally, the transformation procedure usedmay depend upon the host to be transformed. Methods for introducingheterologous polynucleotides into mammalian cells are well known in theart and include, but are not limited to, dextran-mediated transfection,calcium phosphate precipitation, polybrene mediated transfection,protoplast fusion, electroporation, encapsulation of thepolynucleotide(s) in liposomes, and direct microinjection of the DNAinto nuclei.

A nucleic acid molecule encoding the amino acid sequence of a heavychain constant region, a heavy chain variable region, a light chainconstant region, or a light chain variable region of an anti-B7RP1antibody of the invention is inserted into an appropriate expressionvector using standard ligation techniques. In one embodiment, theanti-B7RP1 antibody heavy chain or light chain constant region isappended to the C-terminus of the appropriate variable region and isligated into an expression vector. The vector is typically selected tobe functional in the particular host cell employed (i.e., the vector iscompatible with the host cell machinery such that amplification of thegene and/or expression of the gene can occur). For a review ofexpression vectors, see METHODS IN ENZYMOLOGY 185 (Goeddel, ed.), 1990,Academic Press.

Typically, expression vectors used in any of the host cells will containsequences for plasmid maintenance and for cloning and expression ofexogenous nucleotide sequences. Such sequences, collectively referred toas “flanking sequences” in certain embodiments will typically includeone or more of the following nucleotide sequences: a promoter, one ormore enhancer sequences, an origin of replication, a transcriptionaltermination sequence, a complete intron sequence containing a donor andacceptor splice site, a sequence encoding a leader sequence forpolypeptide secretion, a ribosome binding site, a polyadenylationsequence, a polylinker region for inserting the nucleic acid encodingthe polypeptide to be expressed, and a selectable marker element. Eachof these sequences is discussed below.

Optionally, the vector may contain a “tag”-encoding sequence, i.e., anoligonucleotide molecule located at the 5′ or 3′ end of the anti-B7RP1antibody polypeptide coding sequence; the oligonucleotide sequenceencodes polyHis (such as hexaHis), or another “tag” such as FLAG, HA(hemaglutinin influenza virus), or myc for which commercially availableantibodies exist. This tag is typically fused to the polypeptide uponexpression of the polypeptide, and can serve as a means for affinitypurification or detection of the anti-B7RP1 antibody from the host cell.Affinity purification can be accomplished, for example, by columnchromatography using antibodies against the tag as an affinity matrix.Optionally, the tag can subsequently be removed from the purifiedanti-B7RP1 antibody polypeptide by various means such as using certainpeptidases for cleavage.

Flanking sequences may be homologous (i.e., from the same species and/orstrain as the host cell), heterologous (i.e., from a species other thanthe host cell species or strain), hybrid (i.e., a combination offlanking sequences from more than one source), synthetic or native. Assuch, the source of a flanking sequence may be any prokaryotic oreukaryotic organism, any vertebrate or invertebrate organism, or anyplant, provided that the flanking sequence is functional in, and can beactivated by, the host cell machinery.

Flanking sequences useful in the vectors of this invention may beobtained by any of several methods well known in the art. Typically,flanking sequences useful herein will have been previously identified bymapping and/or by restriction endonuclease digestion and can thus beisolated from the proper tissue source using the appropriate restrictionendonucleases. In some cases, the full nucleotide sequence of a flankingsequence may be known. Here, the flanking sequence may be synthesizedusing the methods described herein for nucleic acid synthesis orcloning.

Whether all or only a portion of the flanking sequence is known, it maybe obtained using polymerase chain reaction (PCR) and/or by screening agenomic library with a suitable probe such as an oligonucleotide and/orflanking sequence fragment from the same or another species. Where theflanking sequence is not known, a fragment of DNA containing a flankingsequence may be isolated from a larger piece of DNA that may contain,for example, a coding sequence or even another gene or genes. Isolationmay be accomplished by restriction endonuclease digestion to produce theproper DNA fragment followed by isolation using agarose gelpurification, Qiagen® column chromatography (Chatsworth, Calif.), orother methods known to the skilled artisan. The selection of suitableenzymes to accomplish this purpose will be readily apparent to one ofordinary skill in the art.

An origin of replication is typically a part of those prokaryoticexpression vectors purchased commercially, and the origin aids in theamplification of the vector in a host cell. If the vector of choice doesnot contain an origin of replication site, one may be chemicallysynthesized based on a known sequence, and ligated into the vector. Forexample, the origin of replication from the plasmid pBR322 (New EnglandBiolabs, Beverly, Mass.) is suitable for most gram-negative bacteria,and various viral origins (e.g., SV40, polyoma, adenovirus, vesicularstomatitus virus (VSV), or papillomaviruses such as HPV or BPV) areuseful for cloning vectors in mammalian cells. Generally, the origin ofreplication component is not needed for mammalian expression vectors(far example, the SV40 origin is often used only because it alsocontains the virus early promoter).

A transcription termination sequence is typically located 3′ to the endof a polypeptide coding region and serves to terminate transcription.Usually, a transcription termination sequence in prokaryotic cells is aG-C rich fragment followed by a poly-T sequence. While the sequence iseasily cloned from a library or even purchased commercially as part of avector, it can also be readily synthesized using methods for nucleicacid synthesis such as those described herein.

A selectable marker gene encodes a protein necessary for the survivaland growth of a host cell grown in a selective culture medium. Typicalselection marker genes encode proteins that (a) confer resistance toantibiotics or other toxins, e.g., ampicillin, tetracycline, orkanamycin for prokaryotic host cells; (b) complement auxotrophicdeficiencies of the cell; or (c) supply critical nutrients not availablefrom complex or defined media. Exemplary selectable markers are thekanamycin resistance gene, the ampicillin resistance gene, and thetetracycline resistance gene. Advantageously, a neomycin resistance genemay also be used for selection in both prokaryotic and eukaryotic hostcells.

Other selectable genes may be used to amplify the gene that will beexpressed. Amplification is the process wherein genes that are requiredfor production of a protein critical for growth or cell survival arereiterated in tandem within the chromosomes of successive generations ofrecombinant cells. Examples of suitable selectable markers for mammaliancells include dihydrofolate reductase (DHFR) and promoterless thymidinekinase genes. Mammalian cell transformants are placed under selectionpressure wherein only the transformants are uniquely adapted to surviveby virtue of the selectable gene present in the vector. Selectionpressure is imposed by culturing the transformed cells under conditionsin which the concentration of selection agent in the medium issuccessively increased, thereby leading to the amplification of both theselectable gene and the DNA that encodes another gene, such as anantibody that binds to B7RP1 polypeptide. As a result, increasedquantities of a polypeptide such as an anti-B7RP1 antibody aresynthesized from the amplified DNA.

A ribosome-binding site is usually necessary for translation initiationof mRNA and is characterized by a Shine-Dalgarno sequence (prokaryotes)or a Kozak sequence (eukaryotes). The element is typically located 3′ tothe promoter and 5′ to the coding sequence of the polypeptide to beexpressed.

In some cases, such as where glycosylation is desired in a eukaryotichost cell expression system, one may manipulate the various pre- orprosequences to improve glycosylation or yield. For example, one mayalter the peptidase cleavage site of a particular signal peptide, or addprosequences, which also may affect glycosylation. The final proteinproduct may have, in the −1 position (relative to the first amino acidof the mature protein) one or more additional amino acids incident toexpression, which may not have been totally removed. For example, thefinal protein product may have one or two amino acid residues found inthe peptidase cleavage site, attached to the amino-terminus.Alternatively, use of some enzyme cleavage sites may result in aslightly truncated form of the desired polypeptide, if the enzyme cutsat such an area within the mature polypeptide.

Expression and cloning vectors of the invention will typically contain apromoter that is recognized by the host organism and operably linked tothe molecule encoding the anti-B7RP1 antibody. Promoters areuntranscribed sequences located upstream (i.e., 5′) to the start codonof a structural gene (generally within about 100 to 1000 bp) thatcontrol transcription of the structural gene. Promoters areconventionally grouped into one of two classes: inducible promoters andconstitutive promoters. Inducible promoters initiate increased levels oftranscription from DNA under their control in response to some change inculture conditions, such as the presence or absence of a nutrient or achange in temperature. Constitutive promoters, on the other hand,uniformly transcribe genes to which they are operably linked, that is,with little or no control over gene expression. A large number ofpromoters, recognized by a variety of potential host cells, are wellknown. A suitable promoter is operably linked to the DNA encoding heavychain or light chain comprising an anti-B7RP1 antibody of the inventionby removing the promoter from the source DNA by restriction enzymedigestion and inserting the desired promoter sequence into the vector.

Suitable promoters for use with yeast hosts are also well known in theart. Yeast enhancers are advantageously used with yeast promoters.Suitable promoters for use with mammalian host cells are well known andinclude, but are not limited to, those obtained from the genomes ofviruses such as polyoma virus, fowlpox virus, adenovirus (such asAdenovirus 2), bovine papilloma virus, avian sarcoma virus,cytomegalovirus, retroviruses, hepatitis-B virus and Simian Virus 40(SV40). Other suitable mammalian promoters include heterologousmammalian promoters, for example, heat-shock promoters and the actinpromoter.

Additional promoters which may be of interest include, but are notlimited to: SV40 early promoter (Bernoist and Chambon, 1981, Nature290:304-10); CMV promoter (Thomsen et al., 1984, Proc. Natl. Acad. Sci.USA 81:659-663); the promoter contained in the 3′ long terminal repeatof Rous sarcoma virus (Yamamoto, et al., 1980, Cell 22:787-97); herpesthymidine kinase promoter (Wagner et al., 1981, Proc. Natl. Acad. Sci.U.S.A. 78:1444-45); promoter and regulatory sequences from themetallothionine gene (Brinster et al., 1982, Nature 296:39-42); andprokaryotic promoters such as the beta-lactamase promoter(Villa-Kamaroff et al., 1978, Proc. Natl. Acad. Sci. U.S.A.,75:3727-31); or the tac promoter (DeBoer et al., 1983, Proc. Natl. Acad.Sci. U.S.A., 80:21-25). Also of interest are the following animaltranscriptional control regions, which exhibit tissue specificity andhave been utilized in transgenic animals: the elastase I gene controlregion that is active in pancreatic acinar cells (Swift et al., 1984,Cell 38:639-46; Ornitz et al., 1986, Cold Spring Harbor Symp. QuantBiol. 50:399-409 (1986); MacDonald, 1987, Hepatology 7:425-515); theinsulin gene control region that is active in pancreatic beta cells(Hanahan, 1985, Nature 315:115-22); the immunoglobulin gene controlregion that is active in lymphoid cells (Grosschedl et al., 1984, Cell38:647-58; Adames et al., 1985, Nature 318:533-38; Alexander et al.,1987, Mol. Cell. Biol., 7:1436-44); the mouse mammary tumor viruscontrol region that is active in testicular, breast, lymphoid and mastcells (Leder et al., 1986, Cell 45:485-95); the albumin gene controlregion that is active in liver (Pinkert et al., 1987, Genes and Devel.1:268-76); the alpha-feto-protein gene control region that is active inliver (Krumlauf et al., 1985, Mol. Cell. Biol 5:16.39-48; Hammer et al.,1987, Science 235:53-58); the alpha 1-antitrypsin gene control regionthat is active in liver (Kelsey et al., 1987, Genes and Devel.1:161-71); the beta-globin gene control region that is active in myeloidcells (Mogram et al., 1985, Nature 315:338-40; Kollias et al., 1986,Cell 46:89-94); the myelin basic protein gene control region that isactive in oligodendrocyte cells in the brain (Readhead et al., 1987,Cell 48:703-12); the myosin light chain-2 gene control region that isactive in skeletal muscle (Sani, 1985, Nature 314:283-86); and thegonadotropic releasing hormone gene control region that is active in thehypothalamus (Mason et al., 1986, Science 234:1372-78).

An enhancer sequence may be inserted into the vector to increasetranscription of DNA encoding light chain or heavy chain comprising ananti-B7RP1 antibody of the invention by higher eukaryotes. Enhancers arecis-acting elements of DNA, usually about 10-300 bp in length, that acton the promoter to increase transcription. Enhancers are relativelyorientation and position independent, having been found at positionsboth 5′ and 3′ to the transcription unit. Several enhancer sequencesavailable from mammalian genes are known (e.g., globin, elastase,albumin, alpha-feto-protein and insulin). Typically, however, anenhancer from a virus is used. The SV40 enhancer, the cytomegalovirusearly promoter enhancer, the polyoma enhancer, and adenovirus enhancersknown in the art are exemplary enhancing elements for the activation ofeukaryotic promoters. While an enhancer may be positioned in the vectoreither 5′ or 3° to a coding sequence, it is typically located at a site5′ from the promoter.

Expression vectors of the invention may be constructed from a startingvector such as a commercially available vector. Such vectors may or maynot contain all of the desired flanking sequences. Where one or more ofthe flanking sequences described herein are not already present in thevector, they may be individually obtained and ligated into the vector.Methods used for obtaining each of the flanking sequences are well knownto one skilled in the art.

After the vector has been constructed and a nucleic acid moleculeencoding light chain, a heavy chain, or a light chain and a heavy chaincomprising an anti-B7RP1 antibody has been inserted into the proper siteof the vector, the completed vector may be inserted into a suitable hostcell for amplification and/or polypeptide expression. The transformationof an expression vector for an anti-B7RP1 antibody into a selected hostcell may be accomplished by well known methods including transfection,infection, calcium phosphate co-precipitation, electroporation,microinjection, lipofection, DEAF-dextran mediated transfection, orother known techniques. The method selected will in part be a functionof the type of host cell to be used. These methods and other suitablemethods are well known to the skilled artisan, and are set forth, forexample, in Sambrook et al., supra.

A host cell, when cultured under appropriate conditions, synthesizes ananti-B7RP1 antibody that can subsequently be collected from the culturemedium (if the host cell secretes it into the medium) or directly fromthe host cell producing it (if it is not secreted). The selection of anappropriate host cell will depend upon various factors, such as desiredexpression levels, polypeptide modifications that are desirable ornecessary for activity (such as glycosylation or phosphorylation) andease of folding into a biologically active molecule

Mammalian cell lines available as hosts for expression are well known inthe art and include, but are not limited to, immortalized cell linesavailable from the American Type Culture Collection (ATCC), includingbut not limited to Chinese hamster ovary (CHO) cells, HeLa cells, babyhamster kidney (BHK) cells, monkey kidney cells (COS), humanhepatocellular carcinoma cells (e.g., Hep G2), and a number of othercell lines. In certain embodiments, cell lines may be selected throughdetermining which cell lines have high expression levels andconstitutively produce antibodies with B7RP1 binding properties. Inanother embodiment, a cell line from the B cell lineage that does notmake its own antibody but has a capacity to make and secrete aheterologous antibody can be selected.

Antibodies of the invention are useful for detecting B7RP1 in biologicalsamples and identification of cells or tissues that produce B7RP1protein. Antibodies of the invention that specifically bind to B7RP1 maybe useful in treatment of B7RP1 mediated diseases Said antibodies can beused in binding assays to detect B7RP1 and to inhibit B7RP1 from forminga complex with B7RP1 receptors. Said antibodies that bind to B7RP1 andblock interaction with other binding compounds may have therapeutic usein modulating B7RP1 mediated diseases. In certain embodiments,antibodies to B7RP1 may block B7RP1 binding to its receptor, which mayresult in disruption of the B7RP1 induced signal transduction cascade.

The present invention also relates to the use of one or more of theantibodies of the present invention in the manufacture of a medicamentfor the treatment of a disorder or condition caused by increasedexpression of B7RP1 or increased sensitivity to B7RP1 in a patient suchas any one of disorders or conditions disclosed herein.

In certain embodiments, the invention provides pharmaceuticalcompositions comprising a therapeutically effective amount of one or aplurality of the antibodies of the invention together with apharmaceutically acceptable diluent, carrier, solubilizer, emulsifier,preservative and/or adjuvant. Acceptable formulation materials arenontoxic to recipients at the dosages and concentrations employed. Inpreferred embodiments, pharmaceutical compositions comprising atherapeutically effective amount of anti-B7RP1 antibodies are provided.

In certain embodiments, acceptable formulation materials are nontoxic torecipients at the dosages and concentrations employed.

In certain embodiments, the pharmaceutical composition may containformulation materials for modifying, maintaining or preserving, forexample, the pH, osmolarity, viscosity, clarity, color, isotonicity,odor, sterility, stability, rate of dissolution or release, adsorptionor penetration of the composition. In such embodiments, suitableformulation materials include, but are not limited to, amino acids (suchas glycine, glutamine, asparagine, arginine or lysine); antimicrobials;antioxidants (such as ascorbic acid, sodium sulfite or sodiumhydrogen-sulfite); buffers (such as borate, bicarbonate, Tris-HCl,citrates, phosphates or other organic acids); hulking agents (such asmannitol or glycine); chelating agents (such as ethylenediaminetetraacetic acid (EDTA)); complexing agents (such as caffeine,polyvinylpyrrolidone, beta-cyclodextrin orhydroxypropyl-beta-cyclodextrin); fillers; monosaccharides;disaccharides; and other carbohydrates (such as glucose, mannose ordextrins); proteins (such as serum albumin, gelatin or immunoglobulins);coloring, flavoring and diluting agents; emulsifying agents; hydrophilicpolymers (such as polyvinylpyrrolidone); low molecular weightpolypeptides; salt-forming counterions (such as sodium); preservatives(such as benzalkonium chloride, benzoic acid, salicylic acid,thimerosal, phenethyl alcohol, methylparaben, propylparaben,chlorhexidine, sorbic acid or hydrogen peroxide); solvents (such asglycerin, propylene glycol or polyethylene glycol); sugar alcohols (suchas mannitol or sorbitol); suspending agents; surfactants or wettingagents (such as pluronics, PEG, sorbitan esters, polysorbates such aspolysorbate 20, polysorbate 80, triton, tromethamine, lecithin,cholesterol, tyloxapal); stability enhancing agents (such as sucrose orsorbitol); tonicity enhancing agents (such as alkali metal halides, forexample, sodium or potassium chloride, mannitol sorbitol); deliveryvehicles; diluents; excipients and/or pharmaceutical adjuvants. SeeREMINGTON'S PHARMACEUTICAL SCIENCES, 18^(th) Edition, (A. R. Gennaro,ed.), 1990, Mack Publishing Company.

In certain embodiments, the optimal pharmaceutical composition will bedetermined by one skilled in the art depending upon, for example, theintended route of administration, delivery format and desired dosage.See, for example, REMINGTON'S PHARMACEUTICAL SCIENCES, supra. In certainembodiments, such compositions may influence the physical state,stability, rate of in vivo release and rate of in vivo clearance of theantibodies of the invention.

In certain embodiments, the primary vehicle or carrier in apharmaceutical composition may be either aqueous or non-aqueous innature. For example, a suitable vehicle or carrier may be water forinjection, physiological saline solution or artificial cerebrospinalfluid, possibly supplemented with other materials common in compositionsfor parenteral administration. Neutral buffered saline or saline mixedwith serum albumin are further exemplary vehicles. In certainembodiments, pharmaceutical compositions of the present inventioncomprise Tris buffer of about pH 7.0-8.5, or acetate buffer of about pH4.0-5.5, and may further include sorbitol, sucrose, Tween-20 and/or asuitable substitute therefor. In certain embodiments of the invention,anti-B7RP1 antibody compositions may be prepared for storage by mixingthe selected composition having the desired degree of purity withoptional formulation agents (REMINGTON'S PHARMACEUTICAL SCIENCES, supra)in the form of a lyophilized cake or an aqueous solution. Further, incertain embodiments, the anti-B7RP1 antibody product may be formulatedas a lyophilizate using appropriate excipients such as sucrose.

The pharmaceutical compositions of the invention can be selected forparenteral delivery. Alternatively, the compositions may be selected forinhalation or for delivery through the digestive tract, such as orally.Preparation of such pharmaceutically acceptable compositions is withinthe skill of the art.

The formulation components are present in concentrations that areacceptable to the site of administration. In certain embodiments,buffers are used to maintain the composition at physiological pH or at aslightly lower pH, typically within a pH range of from about 5 to about8.

When parenteral administration is contemplated, the therapeuticcompositions for use in this invention may be provided in the form of apyrogen-free, parenterally acceptable aqueous solution comprising thedesired anti-B7RP1 antibody in a pharmaceutically acceptable vehicle. Aparticularly suitable vehicle for parenteral injection is steriledistilled water in which the anti-B7RP1 antibody is formulated as asterile, isotonic solution, properly preserved. In certain embodiments,the preparation can involve the formulation of the desired molecule withan agent, such as injectable microspheres, bio-erodible particles,polymeric compounds (such as polylactic acid or polyglycolic acid),beads or liposomes, that may provide controlled or sustained release ofthe product which can be delivered via depot injection. In certainembodiments, hyaluronic acid may also be used, having the effect ofpromoting sustained duration in the circulation. In certain embodiments,implantable drug delivery devices may be used to introduce the desiredantibody molecule.

Pharmaceutical compositions of the invention can be formulated forinhalation. In these embodiments, anti-B7RP1 antibodies areadvantageously formulated as a dry, inhalable powder. In certainembodiments, anti-B7RP1 antibody inhalation solutions may also beformulated with a propellant for aerosol delivery. In certainembodiments, solutions may be nebulized. Pulmonary administration andformulation methods therefore are further described in InternationalPatent Application No. PCT/US94/001875, which is incorporated byreference and describes pulmonary delivery of chemically modifiedproteins.

It is also contemplated that formulations can be administered orally.Anti-B7RP1 antibodies that are administered in this fashion can beformulated with or without carriers customarily used in the compoundingof solid dosage forms such as tablets and capsules. In certainembodiments, a capsule may be designed to release the active portion ofthe formulation at the point in the gastrointestinal tract whenbioavailability is maximized and pre-systemic degradation is minimized.Additional agents can be included to facilitate absorption of theanti-B7RP1 antibody. Diluents, flavorings, low melting point waxes,vegetable oils, lubricants, suspending agents, tablet disintegratingagents, and binders may also be employed.

A pharmaceutical composition of the invention is provided to comprise aneffective quantity of one or a plurality of anti-B7RP1 antibodies in amixture with non-toxic excipients that are suitable for the manufactureof tablets. By dissolving the tablets in sterile water, or anotherappropriate vehicle, solutions may be prepared in unit-dose form.Suitable excipients include, but are not limited to, inert diluents,such as calcium carbonate, sodium carbonate or bicarbonate, lactose, orcalcium phosphate; or binding agents, such as starch, gelatin, oracacia; or lubricating agents such as magnesium stearate, stearic acid,or talc.

Additional pharmaceutical compositions will be evident to those skilledin the art, including formulations involving anti-B7RP1 antibodies insustained- or controlled-delivery formulations. Techniques forformulating a variety of other sustained- or controlled-delivery means,such as liposome carriers, bio-erodible microparticles or porous beadsand depot injections, are also known to those skilled in the art. See,for example, International Patent Application No. PCT/US93/00829, whichis incorporated by reference and describes controlled release of porouspolymeric microparticles for delivery of pharmaceutical compositions.Sustained-release preparations may include semipermeable polymermatrices in the form of shaped articles, e.g. films, or microcapsules.Sustained release matrices may include polyesters, hydrogels,polylactides (as disclosed in U.S. Pat. No. 3,773,919 and EuropeanPatent Application Publication No. EP 058481, each of which isincorporated by reference), copolymers of L-glutamic acid and gammaethyl-L-glutamate (Sidman et al., 1983, Biopolymers 22:547-556),poly(2-hydroxyethyl-methacrylate) (Langer et al., 1981, J. Biomed.Mater. Res. 15:167-277 and Langer, 1982, Chem. Tech. 12:98-105),ethylene vinyl acetate (Langer et al., supra) orpoly-D(−)-3-hydroxybutyric acid (European Patent Application PublicationNo. EP 133,988). Sustained release compositions may also includeliposomes that can be prepared by any of several methods known in theart. See e.g., Eppstein et al., 1985, Proc. Natl. Acad. Sci. USA82:3688-3692; European Patent Application Publication Nos. EP 036,676;EP 088,046 and EP 143,949, incorporated by reference.

Pharmaceutical compositions used for in viva administration aretypically provided as sterile preparations. Sterilization can beaccomplished by filtration through sterile filtration membranes. Whenthe composition is lyophilized, sterilization using this method may beconducted either prior to or following lyophilization andreconstitution. Compositions for parenteral administration can be storedin lyophilized form or in a solution. Parenteral compositions generallyare placed into a container having a sterile access port, for example,an intravenous solution bag or vial having a stopper pierceable by ahypodermic injection needle.

Once the pharmaceutical composition has been formulated, it may bestored in sterile vials as a solution, suspension, gel, emulsion, solid,or as a dehydrated or lyophilized powder. Such formulations may bestored either in a ready-to-use form or in a form (e.g., lyophilized)that is reconstituted prior to administration.

The invention also provides kits for producing a single-doseadministration unit. The kits of the invention may each contain both afirst container having a dried protein and a second container having anaqueous formulation. In certain embodiments of this invention, kitscontaining single and multi-chambered pre-filled syringes (e.g., liquidsyringes and lyosyringes) are provided.

The effective amount of an anti-B7RP1 antibody-containing pharmaceuticalcomposition to be employed therapeutically will depend, for example,upon the therapeutic context and objectives. One skilled in the art willappreciate that the appropriate dosage levels for treatment will varydepending, in part, upon the molecule delivered, the indication forwhich the anti-B7RP1 antibody is being used, the route ofadministration, and the size (body weight, body surface or organ size)and/or condition (the age and general health) of the patient. In certainembodiments, the clinician may titer the dosage and modify the route ofadministration to obtain the optimal therapeutic effect. A typicaldosage may range from about 0.1 μg/kg to up to about 30 mg/kg or more,depending on the factors mentioned above. In certain embodiments, thedosage may range from 0.1 μg/kg up to about 30 mg/kg; from 1 μg/kg up toabout 30 mg/kg; or from 5 μg/kg up to about 30 mg/kg.

Dosing frequency will depend upon the pharmacokinetic parameters of theparticular anti-B7RP1 antibody in the formulation used. Typically, aclinician administers the composition until a dosage is reached thatachieves the desired effect. The composition may therefore beadministered as a single dose, or as two or more doses (which may or maynot contain the same amount of the desired molecule) over time, or as acontinuous infusion via an implantation device or catheter. Furtherrefinement of the appropriate dosage is routinely made by those ofordinary skill in the art and is within the ambit of tasks routinelyperformed by them. Appropriate dosages may be ascertained through use ofappropriate dose-response data. In certain embodiments, the antibodiesof the invention can be administered to patients throughout an extendedtime period. Chronic administration of an antibody of the inventionminimizes the adverse immune or allergic response commonly associatedwith antibodies that are raised against a human antigen in a non-humananimal, for example, a non-fully human antibody produced in a non-humanspecies.

The route of administration of the pharmaceutical composition is inaccord with known methods, e.g. orally, through injection byintravenous, intraperitoneal, intracerebral (intra-parenchymal),intracerebroventricular, intramuscular, intra-ocular, intraarterial,intraportal, or intralesional routes; by sustained release systems or byimplantation devices. In certain embodiments, the compositions may beadministered by bolus injection or continuously by infusion, or byimplantation device.

The composition also may be administered locally via implantation of amembrane, sponge or another appropriate material onto which the desiredmolecule has been absorbed or encapsulated. In certain embodiments,where an implantation device is used, the device may be implanted intoany suitable tissue or organ, and delivery of the desired molecule maybe via diffusion, timed-release bolus, or continuous administration.

It also may be desirable to use anti-B7RP1 antibody pharmaceuticalcompositions according to the invention ex vivo. In such instances,cells, tissues or organs that have been removed from the patient areexposed to anti-B7RP1 antibody pharmaceutical compositions after whichthe cells, tissues and/or organs are subsequently implanted back intothe patient.

In particular, anti-B7RP1 antibodies can be delivered by implantingcertain cells that have been genetically engineered, using methods suchas those described herein, to express and secrete the polypeptide. Incertain embodiments, such cells may be animal or human cells, and may beautologous, heterologous, or xenogeneic. In certain embodiments, thecells may be immortalized. In other embodiments, in order to decreasethe chance of an immunological response, the cells may be encapsulatedto avoid infiltration of surrounding tissues. In further embodiments,the encapsulation materials are typically biocompatible, semi-permeablepolymeric enclosures or membranes that allow the release of the proteinproducts) but prevent the destruction of the cells by the patient'simmune system or by other detrimental factors from the surroundingtissues.

EXAMPLES

The following examples, including the experiments conducted and resultsachieved are provided for illustrative purposes only and are not to beconstrued as limiting the invention.

Example 1 Production of Human Monoclonal Antibodies Against B7 RelatedProtein-1 (B7RP1) Antigen

Purified recombinant human B7RP-1 (hB7RP-1) prepared as described inInternational Patent Application Publication No. WO 00/46240, which isincorporated herein by reference, or CHO cells transfected to expresshB7RP-1 were used as the antigen. Mature human B7RP-1 has the amino acidsequence of residues X to 302 in the sequence shown in WO 00/46240 asSEQ ID NO: 17, wherein X can be 19, 20, 21, 22, 24 or 28.

Transgenic HuMab Mice

Fully human monoclonal antibodies to B7RP-1 were prepared using HCo7 andHCo12 strains of HuMab transgenic mice, both of which express humanantibody genes. In both of these mouse strains, the endogenous mousekappa light chain gene has been homozygously disrupted as described inChen et al (1993) EMBO J. 12:811-820 and the endogenous mouse heavychain gene has been homozygously disrupted as described in Example 1 ofPCT Publication WO 01/09187. Each of these mouse strains carries a humankappa light chain transgene, KCo5, as described in Fishwild et al.(1996) Nature Biotechnology 14:845-851. The HCo7 strain carries the HCo7human heavy chain transgene as described in U.S. Pat. Nos. 5,545,806;5,625,825; and 5,545,807. The HCo12 strain carries the HCo12 human heavychain transgene as described in Example 2 of PCT Publication WO01/09187.

HuMab Immunizations:

To generate fully human monoclonal antibodies to B7RP-1, HuMab mice ofthe HCo7 or HCo12 strain were immunized with purified recombinant B7RP-1or CHO cells transfected to express B7RP-1. General immunization schemesfor HuMab mice are described in Lonberg et al. (1994) Nature 368(6474):856-859; Fishwild et al. (1996) Nature Biotechnology 14: 845-851 and PCTPublication WO 98/24884. The mice were 6-16 weeks of age upon the firstinfusion of antigen. A purified recombinant preparation of B7RP-1antigen (50 μg) or a preparation of transfected CHO cells (3.5×10⁶-1×10⁷cells) was used to immunize the HuMab mice intraperitonealy.

Transgenic mice were immunized twice with purified antigen in completeFreund's adjuvant intraperitonealy, followed by 2-4 weeks of IPimmunizations (up to a total of 8 immunizations) with the purifiedantigen in incomplete Freund's adjuvant. Immunization with CHO cellstransfected to express B7RP-1 was the same except that complete Freund'sadjuvant and incomplete Freund's adjuvant were not used with the cells.The immune response was monitored by retroorbital bleeds. The plasma wasscreened by ELISA (as described below), and mice with sufficient titersof anti-B7RP-1 human immunogolobulin were used for fusions. Mice wereboosted intravenously with antigen 3 and 2 days before sacrifice andremoval of the spleen. Typically, 10-20 fusions for each antigen wereperformed. Several dozen mice were immunized for each antigen. A totalof 28 mice of the HCo7 and HCo12 mice strains were immunized withB7RP-1.

Selection of HuMab Mice Producing Anti-B7RP-1 Antibodies:

To select HuMab mice producing antibodies that bound B7RP-1, sera fromimmunized mice was tested by ELISA as described by Fishwild et al.(1996). Briefly, microtiter plates were coated with purified recombinantB7RP-1 at 1-2 μg/ml in PBS, 50 μl/wells incubated 4° C. overnight thenblocked with 200 μl/well of 5% chicken serum in PBS/Tween (0.05%).Dilutions of plasma from B7RP-1-immunized mice were added to each welland incubated for 1-2 hours at ambient temperature. The plates werewashed with PBS/Tween and then incubated with a goat-anti-human IgG Fcpolyclonal antibody conjugated with horseradish peroxidase (HRP) for 1hour at room temperature. After washing, the plates were developed withABTS substrate (Sigma, A-1888, 0.22 mg/ml) and analyzed byspectrophotometer at OD 415-495. Mice that developed the highest titersof anti-B7RP-1 antibodies were used for fusions. Fusions were performedas described below and hybridoma supernatants were tested foranti-B7RP-1 activity by ELISA.

Generation of Hybridomas Producing Human Monoclonal Antibodies toB7RP-1:

The mouse splenocytes, isolated from the HuMab mice, were fused with PEGto a mouse myeloma cell line based upon standard protocols. Theresulting hybridomas were then screened for the production ofantigen-specific antibodies. Single cell suspensions of spleniclymphocytes from immunized mice were fused to one-fourth the number ofSP2/0 nonsecreting mouse myeloma cells (ATCC, CRL 1581) with 50% PEG(Sigma). Cells were plated at approximately 1×10⁵/well in flat bottommicrotiter plate, followed by about two week incubation in selectivemedium containing 10% fetal bovine serum, 10% P388D1 (ATCC, CRL TIB-63)conditioned medium, 3-5% origen (IGEN) in DMEM (Mediatech, CRL 10013,with high glucose, L-glutamine and sodium pyruvate) plus 5 mM HEPES,0.055 mM 2-mercaptoethanol, 50 mg/ml gentamycin and 1×HAT (Sigma, CRLP-7185). After 1-2 weeks, cells were cultured in medium in which the HATwas replaced with HT. Individual wells were then screened by ELISA(described above) for human anti-B7RP-1 monoclonal IgG antibodies. Onceextensive hybridoma growth occurred, medium was monitored usually after10-14 days. The antibody secreting hybridomas were replated, screenedagain and, if still positive for human IgG, anti-B7RP-1 monoclonalantibodies were subcloned at least twice by limiting dilution. Thestable subclones were then cultured in vitro to generate small amountsof antibody in tissue culture medium for further characterization.

Example 2 Cloning the Anti-B7RP1 Antibody Heavy and Light Chains

The hybridoma expressing the B7RP1 binding monoclonal antibody 16H wasused as a source to isolate total RNA using TRIzol® reagent(Invitrogen). A 5′ RACE (rapid amplification of cDNA ends)oligonucleotide (5′-CGA CUG GAG CAC GAG GAC ACU GAC AUG GAC UGA AGO AGUAGA AA-3′; SEQ ID NO: 69) was ligated to the RNA using the GeneRacer™Kit (Invitrogen) components and protocol. First strand cDNA wassynthesized using a random primer with an extension adapter (5′-GGC CGGATA GGC CTC CAN NNN NNT-3′) (SEQ ID NO: 59) and a 5′ RACE (rapidamplification of cDNA ends) preparative assay was performed using theGeneRacer™ Kit (Invitrogen) according to instructions from themanufacturer. For preparing complete light chain encoding cDNA, theforward primer was the GeneRacer™ nested primer, and the reverse primerwas (5′-GGG GTC AGG CTG GAA CTG AGG-3′) (SEQ ID NO: 60). For preparingcDNA encoding the variable region of the heavy chain, the forward primerwas the GeneRacer™ nested primer and the reverse primer was (5′-TGA GGACGC TGA CCA CAC G-3′) (SEQ ID NO: 61). RACE products were cloned intopCR4-TOPO (Invitrogen) and the sequences determined. Consensus sequenceswere used to design primers for full-length antibody chain PCRamplification.

For preparing cDNA encoding anti-B7RP1 16H kappa light chain, the 5′ PCRprimer encoded the amino terminus of the signal sequence, an XbaIrestriction enzyme site, and an optimized Kozak sequence (5′-CAG CAG AAGCTT CTA GAC CAC CAT GGA CAT GAG GGT CCT CGC TCA GCT CCT GGG-3′) (SEQ IDNO: 62). The 3′ primer encoded the carboxyl terminus and terminationcodon, as well as a SalI restriction site (5′-CTT GTC GAC TCA ACA CTCTCC CCT OTT GAA OCT C-3′) (SEQ ID NO: 63). The resulting PCR productfragment was purified, digested with XbaI and SalI, and then gelisolated and ligated into the mammalian expression vector pDSRα20 (seeInternational Application, Publication No. WO 90/14363, which is hereinincorporated by reference for any purpose pDSRα20 was produced bychanging nucleotide 2563 in pDSRα19 from a “Guanosine” to an “Adenosine”by site directed mutagenesis.).

For preparing cDNA encoding anti-B7RP1 16H heavy chain the 5′ PCR primerencoded the amino terminus of the signal sequence, an XbaI restrictionenzyme site, and an optimized Kozak sequence (5′-ACA ACA AAG CTT CTA GACCAC CAT GGA GTT GGG GCT GAA CTG 0-3′) (SEQ ID NO: 64). The 3′ primerencoded the carboxyl end of the variable region, including a naturallyoccurring sense strand BsmBI site (5′-GTG GAG GCA CTA GAG ACG GTG ACCAGG ATT CC 3′; SEQ ID NO: 65). The resulting product was purified,digested with XbaI and BsmBI, gel isolated and ligated into the pDSRα20vector containing the human IgG1 constant region and also into thepDSRα20 vector containing the human IgG2 constant region. All of thehybridoma derived anti-B7RP1 heavy chain variable regions, regardless ofthe native constant region associated, were cloned as described aboveinto both the pDSRα20 vectors containing the human IgG1 and the humanIgG2 constant regions.

Example 3 Expression of Anti-B7RP1 Antibodies in Chinese Hamster Ovary(CHO) Cells

Stable expression of the 16H anti-B7RP1 mAb was achieved byco-transfection of 16H-heavy chain/pDSRα19 IgG2 B7RP1-kappa/pDSRα19plasmids into dihydrofolate reductase deficient (DHFR⁻) serum-freeadapted Chinese hamster ovary (CHO) cells using a calcium phosphatemethod (the full length 16H heavy chain sequence is shown in SEQ ID NO:44; the 16H kappa chain sequence is shown in SEQ ID NO: 45). Transfectedcells were selected in medium containing dialyzed serum but notcontaining hypoxanthine-thymidine to ensure the growth of cellsexpressing the DHFR enzyme. Transfected clones were screened usingassays such as ELISA in order to detect the expression of 16H anti-B7RP1mAb in the conditioned medium. The highest expressing clones weresubjected to increasing concentrations of methotrexate (MTX) for DHFRamplification. MTX amplified clones were screened using assays such asELISA in order to detect higher expression of 16H anti-B7RP1 mAb in theconditioned medium. The highest expressing clones were subjected tosubcloning to obtain a homogeneous population and creation of cellbanks.

Other recombinant anti-B7RP1 antibodies of the invention can begenerated in Chinese hamster ovary cells deficient in DHFR using thesame protocol as described above for the anti-B7RP1 monoclonal antibody.The DNA sequences encoding the complete heavy chain or light chain ofeach anti-B7RP1 antibody of the invention are cloned into expressionvectors. CHOd-cells are co-transfected with an expression vector capableof expressing a complete heavy chain and an expression vector expressingthe complete light chain of the appropriate anti-B7RP1 antibody. Forexample, to generate a 5D anti-B7RP1 antibody, cells are co-transfectedwith a vector capable of expressing a complete heavy chain comprisingthe amino acid sequence as set forth in SEQ ID NO: 47 and a vectorcapable of expressing a complete light chain comprising the amino acidsequence set forth in SEQ ID NO: 48. Table 2 summarizes exemplarycomplete light chains and exemplary complete heavy chains for anti-B7RP1antibodies having human IgG heavy chain constant regions. One of skillin the art will recognize that the IgG1 or IgG2 could be substituted foreach other (i.e. where IgG1 is listed in the table, IgG2 could bepresent, and vice versa). Alternatively, any other immunoglobulin (e.g.,IgM, IgA, IgE or IgH) could be used to generate antibodies of theinvention.

TABLE 2 Heavy Chain Variable Region + Complete Heavy Antibody HeavyChain Constant Region Chain 16H(IgG2) SEQ ID NO: 7 + SEQ ID NO: 41 SEQID NO: 44 16H(IgG1) SEQ ID NO: 7 + SEQ ID NO: 42 SEQ ID NO: 7016Hg(IgG2) SEQ ID NO: 8 + SEQ ID NO: 41 SEQ ID NO: 46 16Hg(IgG1) SEQ IDNO: 8 + SEQ ID NO: 42 SEQ ID NO: 71 5D(IgG1) SEQ ID NO: 9 + SEQ ID NO:42 SEQ ID NO: 47 5D(IgG2) SEQ ID NO: 9 + SEQ ID NO: 41 SEQ ID NO: 722H(IgG2) SEQ ID NO: 10 + SEQ ID NO: 41 SEQ ID NO: 49 2H(IgG1) SEQ ID NO:10 + SEQ ID NO: 42 SEQ ID NO: 73 2Hg(IgG2) SEQ ID NO: 11 + SEQ ID NO: 41SEQ ID NO: 51 2Hg(IgG1) SEQ ID NO: 11 + SEQ ID NO: 42 SEQ ID NO: 7443H(IgG2) SEQ ID NO: 14 + SEQ ID NO: 41 SEQ ID NO: 52 43H(IgG1) SEQ IDNO: 14 + SEQ ID NO: 42 SEQ ID NO: 75 41H(IgG2) SEQ ID NO: 13 + SEQ IDNO: 41 SEQ ID NO: 54 41H(IgG1) SEQ ID NO: 13 + SEQ ID NO: 42 SEQ ID NO:76 15H(IgG2) SEQ ID NO: 12 + SEQ ID NO: 41 SEQ ID NO: 56 15H(IgG1) SEQID NO: 12 + SEQ ID NO: 42 SEQ ID NO: 57 Light Chain Variable Region +Antibody Light Chain Constant Region Complete Light Chain 16H SEQ ID NO:1 + SEQ ID NO: 43 SEQ ID NO: 45 5D SEQ ID NO: 2 + SEQ ID NO: 43 SEQ IDNO: 48 2H SEQ ID NO: 3 + SEQ ID NO: 43 SEQ ID NO: 50 43H SEQ ID NO: 6 +SEQ ID NO: 43 SEQ ID NO: 53 41H SEQ ID NO: 5 + SEQ ID NO: 43 SEQ ID NO:55 15H SEQ ID NO: 4 + SEQ ID NO: 43 SEQ ID NO: 58

Example 4 Production of Anti-B7RP1 Antibody

Anti-B7RP1 antibody is produced by expression in a clonal line of CHOcells. For each production run, cells from a single vial are thawed intoscrum-free cell culture media. The cells are grown initially in aT-flask followed by spinner flasks and then grown in stainless steelreactors of increasing scale up to a 2000 L bioreactor. Production iscarried out in a 2000 L bioreactor using a fed batch culture, in which anutrient feed containing concentrated media components is added tomaintain cell growth and culture viability. Production lasts forapproximately two weeks during which time anti-B7RP1 antibody isconstitutively produced by the cells and secreted into the cell culturemedium.

The production reactor is controlled at a predetermined pH, temperature,and dissolved oxygen level: pH is controlled by carbon dioxide gas andsodium carbonate addition; dissolved oxygen is controlled by air,nitrogen, and oxygen gas flows.

At the end of production, the cell broth is fed into a disk stackcentrifuge and the culture supernatant is separated from the cells. Theconcentrate is further clarified through a depth filter followed by a0.2 μm filter. The clarified conditioned media is then concentrated bytangential flow ultrafiltration. The conditioned media is concentrated15- to 30-fold. The resulting concentrated conditioned medium is theneither processed through purification or frozen for purification at alater date.

Example 5 Germlining the 16H mAb

Sequence alignment of the 16H antibody with human germline sequencesshowed that the framework sequence in the variable region of the 16Hantibody was most identical to the V_(H) 3-07 and JH4 germlinesequences, with only three amino acid differences (FIG. 1A). Theframework sequence for the VK region of the 16H antibody was found to beidentical to the VK1-L15 germline sequence. It is theoretically possiblethat somatic hypermutations are recognized as foreign by the immuneresponse of a patient; in which case the patient would generate ananti-idiotype response that could neutralize the therapeutic. To reducethis possibility, the three amino acid changes in the VH frameworkregion were converted back to the V_(H) 3-07 and JH4 germline sequences(FIG. 1A). Since the germline V_(H) and J_(H) gene segments are presentin every human genome, the germline version of 16H is not likely to berecognized as foreign by the immune response of a dosed patient. Plateco-stimulation bioassays were conducted to determine if the germlinedantibodies could induce T-cell proliferation with an IC₅₀ similar to theIC₅₀ of the non-germlined antibodies. The co-stimulation assays wereconducted as described below using anti-CD3 and hB7RP-1-Fc fusionprotein confirmed that this germlined antibody, referred to as16Hgermline or 16Hg, retains its biological activities (FIG. 1B).

Example 6 Affinity Measurement of Monoclonal Antibodies by Biacore® andKinExA

Three antibodies (5D and 16H, prepared as described in Example 1, and16H germline, prepared as described in Example 5) were purified andsubmitted to binding affinity analysis. B7RP1-Fc was immobilized at ahigh density on a CMS sensor chip using standard amine couplingchemistry. A fixed concentration of mAb was then incubated with varyingconcentrations of B7RP-1 or B7RP1-Fc for at least eight hours at roomtemperature to allow them to reach equilibrium. The samples were theninjected over the B7RP1-Fc surface, and the binding signal observedrepresented free antibody remaining in solution at equilibrium. By usingtwo different antibody concentration (0.2 nM and 1 nM), the K_(D) of theinteraction between a particular mAb and ligand was calculated fromnonlinear regression analysis of the competition curves using adual-curve one-site homogeneous binding model (Adamczyk et al., 1999,Bioconjugate Chem. 10:10.32-37; Adamczyk et al., 2000, Methods20:319-28). As shown in FIG. 2 and Table 3, the 16H, 16Hg, and 5D mAbsall bound both soluble B7RP-1 and B7RP-1-Fc proteins at high affinities.In addition, the results indicated that the 16H (non-germline) and the16Hg (germline) reacted similarly, demonstrating that germlining did notsignificantly affect binding between antibody and ligand.

TABLE 3 Summary of K_(D) Values B7RP-1 B7RP1-Fc  5D  37 pM 1.6 pM  16H1.9 nM 27 pM 16H (germline) 2.7 nM 17 pM

Binding of 5D, 21-1, and 2H germline antibodies was also tested usingKinExA (kinetic exclusion assay) technology. In this assay, hB7RP-1 wascoupled to agarose beads. The beads were used to create a bead column.Samples containing antibody at a fixed concentration, which were allowedto come to equilibrium with varying concentrations of hB7RP-1, were thenpassed over the bead column. Antibody not complexed with ligand bound tothe coated beads.

A fluorescent tagged anti-human Fc secondary antibody was used to detectbound test antibody. The signal obtained was proportional to freeantibody in solution at a given ligand concentration. Using twodifferent antibody concentrations, the K_(D) of the interaction wascalculated from nonlinear regression analysis of the competition curvesusing a dual-curve one-site homogeneous binding model (Adamczyk et al.,1999, Bioconjugate Chem. 10:1032-37; Adamczyk et al., 2000, Methods20:319-28). FIGS. 3, 4, and 5 show the dual-curve fits for antibodies5D, 2H and 2H (germline). Using this technique, an approximately 10-folddifference was seen in the K_(D)s for antibodies 5D and 2H.

The results of the Biacore® and KinExA assays demonstrated that antibody5D has a higher affinity for hB7RP-1 than do either 2H or 16H. Also, thegermline version of antibody 2H does not show a significant differencefrom the non-germline construct.

Example 7 Functional Characteristics of anti-B7RP1 Antibodies

The functional characteristics of B7RP-1 antibodies of the inventionwere evaluated using binding-competition assays, in vitro co-stimulationassays and in vitro tetanus toxoid assays.

Binding-Competition Studies

Binding-competition studies were conducted with the 16H mAbs todemonstrate that they can compete for ICOS binding for B7RP-1. CHO cellstransfected with a gene encoding the full-length human B7RP-1 were firstincubated with decreasing amounts of unlabeled 16H mAb and subsequentlystained with a fluorescently-labeled ICOS-Fc fusion protein. The cellswere then analyzed using flow cytometry. As shown in FIG. 6, ICOS-Fcstained the B7RP-1-transfected CHO cells; 0.4 μg/ml of 16H mAb did notaffect ICOS-Fc binding. However, 6 and 25 μg/ml of 16H efficientlycompeted away ICOS-Fc binding, indicating that the 16H mAb indeedcompeted for ICOS binding on B7RP-1.

Co-Stimulation Assays

Cell culture plates (Falcon, Cat NO_(—)353077, U bottom) were coatedwith 1 μg/ml anti-human CD3 antibodies (PharMingen Cat No. 555336) and10 ug/ml anti-human IgG (Fc specific, Sigma Cat No. I3391). The anti-CD3antibodies and anti-human immunoglobulin in phosphate buffered saline(PBS) were added to each well (100 μl/well) The coated plates wereincubated at 4° C. overnight or at room temperature for 2 hours. Theplates were then washed with PBS twice. After washing, 1 μg/ml humanB7-2Fc (R&D System, Cat No. 141-B2) or 5 μg/ml hB7RP1Fc, each diluted inPBS, were added to each well (100 μl per well). The plates were thenincubated at room temperature for 3 hours and washed twice with PBSthereafter. Purified human T cells were added (1×10⁵ per well) in 200 μlvolume of media (RPMI 1640 supplemented with 10% fetal calf serum (FCS),penicillin-streptomycin-L-glutamine (PSG), β-mercpatoethanol (2-ME),N-Acetyl aspartate (NAA) and Napyruvate) and incubated at 37° C., 5% CO₂for 48 hours. 3H thymidine (ICN Cat No. 2404205) was added at 1 μCi/welland the cells were incubated overnight at 37° C., 5% CO₂. The cells werethen harvested and counted.

Cell culture plates (Falcon, Cat No. 353077, U bottom) were coated with0.1 μg/ml anti-human CD3 as above. hB7RP1 transfected CHO cells (5000RADirradiated) were added at 2×10⁴ per well followed by purified human Tcells at 1×10⁵ per well in 2000 volume. Plates were incubated at 37° C.,5% CO₂ for 48 hours as above. ³H thymidine was added at 1 μCi/well Cellswere incubated overnight, harvested, and counted as above.

Tetanus Toxoid Assays

PBMC were purified from human blood using a Ficoll-Paque (AmershamBiosciences) gradient as follows. Blood was diluted 1:2 with PBS,diluted blood was layered on top of the Ficoll (⅓ room temp Ficoll+⅔diluted blood), centrifuged at 2500 rpm for 30 minutes at roomtemperature, the top layer (plasma & platelets) was aspirated off, andthe mononuclear cell layer was transferred to a fresh 50 ml tube. Theisolated PBMC were washed with PBS (3× the volume of the mononuclearcell layer) and centrifuged for 10 minutes at 1300 rpm at roomtemperature and washed as above. The PBMC were resuspended in media(RPMI 1640+10% heat-inactivated FBS+1× PSG+1× NEAA+55 μM 2-ME) and thecells were counted.

PMBC were added to wells of a 96-well round bottom plate at 100PBMC/well (3×10⁶/ml). Tetanus toxoid (20 μg/ml; University ofMassachusetts) was added for a final concentration of 5 μg/ml. The cellswere incubated for 3 days at 37° C.; 100 μl supernatant were collectedand incubated for an addition 6 to 8 hours in the presence of 1 μCi/well³H-thymidine (MP Biomedicals). The cells were then harvested andcounted.

Table 4 summarizes the functional characteristics of certain antibodiesof the invention as determined using the assays described above.

TABLE 4 Biacore Plate Biacore Fc mono CHO Tetanus Toxoid  2H 43 89 144515 15H 36 141 16H 53 27 1900 276 27 16Hg 32 17 2700 523 41H 52 115 43H46 35  5D 55 1.6 37 1456 15 ICOS-Fc 200-1000 1000 10,672 α-CD86 40*EC₅₀/KD values in pM

Example 8 Epitope Mapping

Experiments were conducted to identify the region on B7RP-1 to which the16H/16Hg and 5D monoclonal antibodies bind. To do this, a novelFluorescence-Activated Cell Sorter (FACS) binding assay was developed.The human extracellular domain (ECD) of B7RP1 (SEQ ID NO: 66) as well astruncated forms of B7RP-1 containing either the Ig1 (IgV-Iike; SEQ IDNO: 67) or the Ig2 (IgC-like; SEQ ID NO: 68) were expressed asN-terminal, in-frame fusions with chicken avidin,

SEQ ID NO: 66 (ECD): DTQEKEVRAMVGSDVELSCACPEGSRFDLNDVYVYWQTSESKTVVTYHIPQNSSLENVDSRYRNRALMSPAGMLRGDFSLRLFNVTPQDEQKFHCLVLSQSLGFQEVLSVEVTLHVAANFSVPVVSAPHSPSQDELTFTCTSINGYPRPNVYWINKTDNSLLDQALQNDTVFLNMRPGLYDVVSVLRIARTPSVNIGCCIENVLLQQNLIVGSQTGNDIGERDKITENP SEQ ID NO: 67 (IgV-like):DTQEKEVRAMVGSDVELSCACPEGSRFDLNDVYVYWQTSESKTVVTYHIPQNSSLENDSRYRNRALMSPAGMLRGDFSLRLFNVTPQDEQKFHCLVLSQSLGFQEVLSVEVTLHVAANFSVPVVSAPHSPSQDELTFT SEQ ID NO: 68 (IgC-like):LGFQEVLSVEVTLHVAANFSVPVVSAPHSPSQDELTFTCTSINGYPRPNVYWINKTDNSLLDQALQNDTVFLNMRGLYDVVSVLRIARTPSVNIGCCIENVLLQQNLTVGSQTGNDIGERDKITENP

Expression vectors containing genes encoding these fusion proteins wereindividually transiently transfected into 293T cells and the conditionedmedia from these cell lines were used as the source of fusion protein.The avidin-tag was used to capture the B7RP1 fusion proteins fromsolution using a biotincoated bead. Fusion proteins were incubated witheither fluorescently-labeled 16H or 5D mAbs or a fluorescently-labeledICOS-Fc fusion protein, and incubated with biotin-coated beads The beadswere recovered and analyzed using flow cytometry on a Becton-DickinsonBioscience FACScan (BD, Franklin Lakes, N.J.). As shown in FIG. 9A,fluorescent staining of the beads was detected with the 16H, 5D, and theICOS reagents when the full ECD of B7RP-1 was attached, indicating thatall three of these reagents could bind to the ECD of B7RP-1. Similarly,all three reagents bound to the avidin fusion protein containing onlythe Ig1 domain, indicating that both ICOS and the blocking anti-B7RP-1mAbs could bind to this region. In contrast, neither ICOS nor theanti-B7RP-1 mAbs could bind to the fusion protein containing only themembrane-proximal Ig2 domain. Thus, the ICOS, 16H, and 5D bindingregions on B7RP-1 were located in the Ig1 domain.

The antibodies generated as described above in Example 1 and tested forbinding using the avidin fusion binding assay, could be divided into twoepitope classes, H and D, as shown in Table 5. Of the 100 antibodiesinitially selected based on their ability to bind B7RP1, 15 failed tobind in the avidin fusion binding assay, most likely because ofdegradation.

TABLE 5 Classification of mAbs by epitope Class # H epitope 75 D epitope10 New epitope, ICOS blocker 0 New epitope, not an ICOS blocker 0 Nodetectable binding 15

Example 9 SNP Identification and Functional Analysis

One major single nucleotide polymorphism (SNP) variant was identified inB7RP-1 that is present in the population with an allele frequency of28.4% (FIG. 7). The variant was identified within the mature proteincoding sequence. A search of the National Center for BiotechnologyInformation (NCBI) databank revealed a second potential SNP variant; thesecond variant was identified in a 1.5 individual (three chromosome)analysis. The first SNP variant (V128I) was located in the firstIgV-like domain, whereas the NCBI SNP variant (L221F) was located in thesecond IgC-like domain.

As discussed above, both the 16H and 5D monoclonal antibodies bind tothe first IgV-like domain, this it is unlikely that the latter L221Fvariant affects either 161-1 or 5D mAb binding or function. Nonetheless,to determine if either of these SNP variants affects 16H or 5D bindingand/or function, two different experiments were conducted. In the firstset of experiments, avidin fusion proteins were constructed with the twoSNP variants and tested for binding to 16H or 5D antibodies in the flowcytometric assay as described above. These representative mAbs from theH and D epitope classes bound to the SNP variants with similar efficacyas the wild-type B7RP-1 (FIG. 9B). These data suggested that antibodiesfrom both the H and D epitope classes bind to the B7RP-1 SNP variants.

In the second approach, Fc fusion proteins were constructed using theB7RP-1 SNP variant sequences and compared for the ability of theseproteins to stimulate T cells in the plate co-stimulation assay (FIG.9C). Both the 16H and 5D antibodies inhibited co-stimulation mediated bythe SNP variant Fc fusion proteins with similar EC₅₀s as the wild-typefusion protein. Taken together these data indicated that the twopotential B7RP-1 SNP variants were recognized by the antibodies of theinvention. Thus, the antibodies of the invention can bind to target inpatients containing these SNP variants.

Example 10 In Vivo Animal Efficacy Models

The ability of B7RP-1 antibodies to inhibit immune response was analyzedusing a murinized rat anti-murine B7RP-1 monoclonal antibody (1B7v2) andchallenging BALB/c mice with keyhole-limpet hemocyanin (KLH).

Generation of the Murinized Rat Anti-Murine B7RP-1 Monoclonal Antibody1B7v2

A Chinese-Hamster-Ovary cell line that overexpressed a full-lengthmurine B7RP-1 was injected into rats as a primary immunization, andsubsequently with a murine B7RF-1-Fc fusion protein to boost the immuneresponse. Spleens were harvested 3 or 4 days post-intravenous boost andthe splenic B cells fused with the Y3-Ag1.2.3 rat myeloma line (ATCCCRL-1631). Cells were then selected in media supplemented withhypoxanthine-aminopterin-thymidine (HAT) for 2 weeks and subsequentlysingle-cell subcloned by limiting dilution. These procedures aredescribed in “Practical Immunology, 2nd ed.” Leslie Hudson and Frank C.Hay; Blackwell Scientific Publications 1980.

Genes encoding the 1B7 immunoglobulin were cloned from the 1B7 cell lineusing standard procedures (Sambrook et al., 2001, MOLECULAR CLONING: ALABORATORY MANUAL, 3d ed., Cold Spring Harbor Laboratory Press, ColdSpring Harbor, N.Y). The isotype switch of the human anti-huB7RP1 MAbswas accomplished by cloning the variable region fragments containingXbaI and BsmBI restriction site cohesive ends into the pDSRa vector withthe human IgG1 or huIgG2 constant region which also had XbaI and BsmBIends. For the 1B7 rat anti-muB7RP1 the chimera was formed by a threestep overlapping PCR process. The rat variable region was PCR amplifiedwith a 3′ primer that contained part, ˜25-35 nucleotides, of the murineconstant region. The murine constant region was amplified with a 5′primer that contained part, ˜25-35 nucleotides, of the rat variableregion. The two fragments were then used as template and the 5′ ratvariable region Mal containing) and the murine 3′ constant region (SalIcontaining) primers were used to generate a complete light chain orheavy chain. The light chain and heavy chain PCR products were thendigested with XbaI and SalI and cloned into pDSRα19. A total of 25 μg oflinearized DNA (12.5 μg pDC323B LC+12.5 μg pDC324 HC) were transfectedinto CS-9 cells using electroporation and selected on DHFR-supplementedmedium.

To test the efficacy of the 1B7v2 mAb, plate co-stimulation assays wereconducted with this mAb. The results were compared with otheranti-murine B7RP-1 mAbs (FIG. 10A) As discussed above, 1B7 is theoriginal hybridoma-produced mAb; two different preparations (labeled1.33 and 7A) were tested. 5E1 and 11G10 were other anti-mB7RP-1monoclonals generated in the fusions described above. Finally, HK5.3 wasa commercially-available anti-rnB7RP-1 (ebiosciences # 16-5985-85).

The 1B7v2 mAb blocked T cell activation in this assay equal to or betterthan any of the other mAbs, and thus was selected as the surrogatetherapeutic for further studies.

Antigen Challenge in Mice

Keyhole Limpet Hemocyanin (KLH) was purchased from Pierce Biotechnology(Rockford, Ill.). Dosing solution #1 (KLH 5 mg/kg in 1 mg/mouse ALUM)was prepared with equal parts of 2×ALUM (500 mg of ALUM plus 50 ml PBS(phosphate buffered saline)) and 2×KLH (2.0 ml dH20 (RNAse-Free) mixedwith 20 mg of lyophilized KLH, brought to 20 ml with 1×PBS). Dosingsolution #2 (KLH 1 mg/kg in 1 mg/mouse ALUM) was prepared with 1 part2×KLH mixed with 4 parts 1× phosphate buffered saline.

Female BALB/c mice were primed either with 1 mg/kg of KLH/alum andre-immunized on day 21 with 5 mg/kg KLH only, introduced byintraperitoneal injection. Mice were treated by intraperitonealinjection with 1B7v2, the isotype control antibody (anti-AGP3 PB) or thevehicle (PBS) alone, starting on day 1 (one day prior to priming withKLH/alum) in a final volume of 200 μl every 5 days.

The mice were bled every 7 days retro-orbitally (approximately 200 μl)to obtain approximately 50-100 μl of serum for analysis ofantigen-specific serum IgM (FIG. 10B), IgG2a (FIG. 10C), and IgG1 (FIG.10D) Both the isotype-control and vehicle-treated mice showedsignificant primary and secondary immune responses. The IgM response wasnot affected by treatment, whereas, blockade of B7RP-1-ICOS with 1B7v2decreased both primary and secondary IgG2a and IgG1 responses in astatistically-significant manner.

IL-5 is a cytokine released by T cells in response to antigenstimulation that induces B cell differentiation and function. As theB7RP-1/ICOS interaction is believed to be critical for T-cell-dependentB cell function, measuring serum IL-5 levels was used to determine ifinterdiction of the B7RP-1/ICOS axis was indeed affecting T cellfunction. As expected, blockade of B7RP-1 also inhibited antigen-inducedserum IL-5 levels. Sera were harvested from the mice from the antigenchallenge experiment outlined above 24 hours after the antigen challengeon day 21, and serum IL-5 levels were determined by ELISA. As shown inFIG. 11, elevated IL-5 levels were detected in the test mice as early as9 hours after challenge; levels began to decline by 48 hours andreturned to baseline by 72 hours. Treatment of the mice with 1B7v2 mAblead to a statistically significant repression of IL-5 levels at the24-hour time point.

Example 11 Binding to Cynomolgus Monkey B7RP-1

To determine if the anti-hB7RP-1 mAbs also bind to cynomolgus monkeyB7RP-1, flow cytometric staining experiments were conducted with the 16HmAb and B cells purified from cynomolgus monkeys and humans. As shown inFIG. 12A, addition of fluorescently-labeled 16H to cyno B cells lead tostaining, indicating that 16H was indeed binding to cyno B7RP-1 (rightpanel). As expected, 16H also stained human B cells (left panel). Inaddition, 16H, 16Hg, and 5D were tested in plate co-stimulation assaysusing cyno T cells, cyno B7RP-1-Fc, and anti-CD3 mAb. As shown in FIG.12B, all three mAbs inhibited cyno B7RP-1-dependent cyno T cellactivation, indicating that these mAbs functionally block the cynoICOS-B7RP-1 interaction.

Example 12

T-Cell Dependent Antigen Responses in the Cynomolgus Monkey FollowingAdministration of the Anti-B7RP-1 Antibodies

A cynomolgus monkey study was conducted with two anti-B7RP-1 monoclonalantibodies, 1611 and 5D, to assess the ability of these antibodies toinhibit a T-cell dependent B cell antigen response as determined byserum levels of antigen-specific antibody. Briefly, the anti-keyholelimpet hemocyanin (KLH) and anti-tetanus toxoid antibody responses wereexamined following antigen challenge in the presence of B7RP-1antibodies in the cynomolgus monkey.

Test Article 1 was 1611 and Test Article 2 was 5D. The Control Articlewas the vehicle for B7RP-1 antibody (0.01 sodium acetate, pH 5.0, 5%sorbitol, 0.004% Tween 20). Keyhole Limpet Hemocyanin (KLH) waspurchased from Pierce Biotechnology (Rockford, Ill.).

The KLH was prepared by reconstitution with sterile water to yield a 10mg/mL stock solution. The stock solution was diluted with sterile waterto yield a 1 mg/mL dosing solution. Tetanus Toxoid used for theseexperiments was Super-Tet® Tetanus Toxoid w/Havlogen©, purchased fromIntervet™ Inc. (Milsboro, Del.). The dose level for these experimentswas 75 IU (0.5 mL of 150 IU/mL).

Table 6 shows the treatment group distribution of 28 cynomolgus monkeys

TABLE 6 Dose Number of Dose Dose Solution Group Males/ Level VolumeConc. No. Females Test Article Route (mg/kg) (mL/kg) (mg/mL) 1 2/2Control IV 0 1 0 2 2/2 B7RP-1 5D lV 0.1 1 0.1 3 2/2 B7RP-1 5D IV 1.0 11.0 4 2/2 B7RP-1 5D IV 8.0 1 10.0 5 2/2 B7RP-1 16H lV 0.1 1 0.1 6 2/2B7RP-1 16H IV 1.0 1 1.0 7 2/2 B7RP-1 16H IV 8.0 1 10.0

Test article doses were administered via intravenous injection to allanimals on Days 1, 8, 15, 22, 29, 36, 43, and 50. Animals scheduled fornecropsy in Groups 1-4 (1/sex/group) received an additional dose on Day57. Evaluation of immune response was conducted on all animals viaimmunization with KLH and tetanus toxoid antigens followed by bloodsampling for antigen-specific immunoglobulins (IgM and IgG).

Titer values were present following primary administration of both theKLH and tetanus antigens. For KLH, primary titer values ranged from 0 to900 for both IgM and IgG. As the primary KLH challenge was administeredprior to test article administration, no effect of the B7RP-1 antibodieswas evaluated. For tetanus toxoid, primary titer values ranged from 0 to50 for IgM and from 0 to 4050 for IgG. There were no differences in theprimary response to tetanus toxoid between the B7RP-1 antibody groupsand the control group.

As expected, titer values for IgG were increased following secondaryadministration of both the KLH and tetanus antigens, when compared tothe primary titer values. For KLH, secondary titer values ranged from 0to 300 for IgM and from 0 to 8100 for IgG. However, there was noevidence of inhibition of the KLH secondary response attributed toadministration of the B7RP-1 antibodies.

For tetanus toxoid, secondary titer values were below 50 for IgM andranged from 1350 to 36450 for IgG. Results for individual animal andgroup mean values are presented in the FIG. 13A (16H antibody) and FIG.13B (5D antibody) for Days 53 and 57 following the secondary challengewith tetanus toxoid on Day 42.

On Day 53, the number of animals reaching peak response was 3/4, 1/4,and 1/4 at the 0.1, 1, and 8 mg/kg dose levels of 16H, respectively, and1/4, 1/4, and 1/4 at the 0.1, 1, and 8 mg/kg dose levels of 5Drespectively, compared to 214 control animals. Thus, in general, thenumber of animals reaching a high titer on Day 53 was reduced in theB7RP-1 antibody-treated groups. On Day 57, titer values were maintainedin the control animals, while titer values for several of the B7RP-1antibody-treated animals declined from the Day 5.3 values. The number ofanimals with high titers on Day 57 was 0/4, 0/4, and 1/4 at the 0.1, 1,and 8 mg/kg dose levels of 16H, respectively, and 0/4, 0/4, and 0/4 atthe 0.1, 1, and 8 mg/kg dose levels of 5D respectively, compared to 2/4control animals.

These results demonstrated that the two B7RP-1 antibodies 16H and 5Dinhibited a T-cell dependent B cell antigen response in cynomolgusmonkeys, as determined by serum levels of tetanus toxoid-specificantibody. In addition, the presence of the B7RP-1 antibodies wasimportant for blockage of the B7RP-1-ICOS interaction during the primaryresponse in order to detect an effect following the secondary challenge.

These results and the results from Example 10 demonstrated that both thesurrogate therapeutic and the therapeutic candidates blocked T and Bcell-dependent immune responses in murine and monkey model systems,which indicated that blocking this co-stimulatory axis may beefficacious in the treatment of B-cell-mediated diseases such asSystemic Lupus Erythematosus (SLE), asthma, and Rheumatoid Arthritis(RA).

It should be understood that the foregoing disclosure emphasizes certainspecific embodiments of the invention and that all modifications oralternatives equivalent thereto are within the spirit and scope of theinvention as set forth in the appended claims.

1. An isolated nucleic acid molecule that encodes an antibody orantigen-binding fragment thereof, that binds specifically to humanB7RP1, wherein the antibody or antigen-binding fragment thereofcomprises a heavy chain variable region and a light chain variableregion, wherein the heavy chain variable region comprises any one of SEQID NOS: 7-14.
 2. An isolated nucleic acid molecule that encodes anantibody or antigen-binding fragment thereof that binds specifically tohuman B7RP1 comprising: a) a heavy chain having a heavy chain variableregion comprising SEQ ID NO: 7 and a light chain having a light chainvariable region comprising SEQ ID NO: 1; b) a heavy chain having a heavychain variable region comprising SEQ ID NO: 8 and a light chain having alight chain variable region comprising SEQ ID NO: 1; c) a heavy chainhaving a heavy chain variable region comprising SEQ ID NO: 9 and a lightchain having a light chain variable region comprising SEQ ID NO: 2; d) aheavy chain having a heavy chain variable region comprising SEQ ID NO:10 and a light chain having a light chain variable region comprising SEQID NO: 3; e) a heavy chain having a heavy chain variable regioncomprising SEQ ID NO: 11 and a light chain having a light chain variableregion comprising SEQ ID NO: 3; f) a heavy chain having a heavy chainvariable region comprising SEQ ID NO: 12 and a light chain having alight chain variable region comprising SEQ ID NO: 4; g) a heavy chainhaving a heavy chain variable region comprising SEQ ID NO: 13 and alight chain having a light chain variable region comprising SEQ ID NO:5; or h) a heavy chain having a heavy chain variable region comprisingSEQ ID NO: 14 and a light chain having a light chain variable regioncomprising SEQ ID NO:
 6. 3. An isolated nucleic acid molecule thatencodes an antibody or antigen-binding fragment thereof that bindsspecifically to human B7RP1, wherein the antibody or antigen-bindingfragment thereof comprises a heavy chain and a light chain, wherein thelight chain comprises a light chain variable region comprising any oneof SEQ ID NOS: 1-6.
 4. The nucleic acid molecule of claim 2, wherein theheavy chain comprises a heavy chain variable region comprising SEQ IDNO: 8, and the light chain variable region comprises SEQ ID NO:
 1. 5. Anisolated nucleic acid molecule that encodes an antibody orantigen-binding fragment thereof that binds specifically to human B7RP1,wherein the antibody or antigen-binding fragment thereof comprises aheavy chain and a light chain, the heavy chain comprising: a) a heavychain CDR1 comprising SEQ ID NO: 27, a heavy chain CDR2 comprising SEQID NO: 28, and a heavy chain CDR3 comprising SEQ ID NO: 29; b) a heavychain CDR1 comprising SEQ ID NO: 30, a heavy chain CDR2 comprising SEQID NO: 31, and a heavy chain CDR3 comprising SEQ ID NO: 32; c) a heavychain CDR1 comprising SEQ ID NO: 27, a heavy chain CDR2 comprising SEQID NO: 33, and a heavy chain CDR3 comprising SEQ ID NO: 34; d) a heavychain CDR1 comprising SEQ ID NO: 35, a heavy chain CDR2 comprising SEQID NO: 36, and a heavy chain CDR3 comprising SEQ ID NO: 37; e) a heavychain CDR1 comprising SEQ ID NO: 27, a heavy chain CDR2 comprising SEQID NO: 33, and a heavy chain CDR3 comprising SEQ ID NO: 38; or f) aheavy chain CDR1 comprising SEQ ID NO: 35, a heavy chain CDR2 comprisingSEQ ID NO: 39, and a heavy chain CDR3 comprising SEQ ID NO:
 40. 6. Thenucleic acid molecule of claim 5, wherein the heavy chain comprises aheavy chain CDR1 comprising SEQ ID NO: 27, a heavy chain CDR2 comprisingSEQ ID NO: 28, and a heavy chain CDR3 comprising SEQ ID NO:
 29. 7. Thenucleic acid molecule of claim 6, wherein the light chain comprises alight chain CDR1 comprising SEQ ID NO: 15, a light chain CDR2 comprisingSEQ ID NO: 16, and a light chain CDR3 comprising SEQ ID NO:
 17. 8. Thenucleic acid molecule of claim 1, 2, 3, or 5, wherein the antibody orantigen-binding fragment thereof is a single-chain antibody, a singlechain Fv antibody, an Fab antibody, an Fab′ antibody, or an (Fab′)₂antibody.
 9. The nucleic acid molecule of claim 1, 2, 3, or 5, whereinthe antibody or antigen-binding fragment thereof is a fully humanantibody or antigen-binding fragment thereof.
 10. The nucleic acidmolecule of claim 1, 2, 3, or 5, wherein the antibody or antigen-bindingfragment thereof blocks interaction between B7RP1 and ICOS.
 11. A vectorcomprising the nucleic acid of claim 1, 2, 3, or
 5. 12. A host cellcomprising the vector of claim
 11. 13. An isolated nucleic acid moleculecomprising a nucleotide sequence that encodes a peptide comprising anyone of SEQ ID NO: 1-14, 44-58, and 70-76, or a combination thereof. 14.A vector comprising the nucleic acid molecule of claim 13
 15. A hostcell comprising the vector of claim
 14. 16. An isolated nucleic acidmolecule that encodes an antibody or antigen-binding fragment thereofthat binds specifically to human B7RP1, wherein the antibody orantigen-binding fragment thereof binds to a region of B7RP1 consistingof SEQ ID NO: 67 and does not bind a region of B7RP1 consisting of SEQID NO:
 68. 17. An isolated nucleic acid molecule that encodes anantibody or an antigen-binding fragment thereof that binds to humanB7RP1, wherein the antibody or antigen-binding fragment thereof iscompetitive with an antibody that comprises a light chain variableregion or SEQ ID NO: 1 and a heavy chain variable region of SEQ ID NO: 8for binding to human B7RP1.
 18. The nucleic acid molecule of claim 16 or17, wherein the antibody or antigen-binding fragment thereof is asingle-chain antibody, a single-chain Fv antibody, an Fab antibody, anFab′ antibody, or an (Fab′)₂ antibody.
 19. The nucleic acid molecule ofclaim 16 or 17, wherein the antibody or antigen-binding fragment thereofis a fully human antibody.
 20. The nucleic acid molecule of claim 16 or17, wherein the antibody or antigen-binding fragment thereof blocksinteraction between B7RP1 and ICOS.
 21. A vector comprising the nucleicacid molecule of claim 24 or
 25. 22. A host cell comprising the vectorof claim
 32. 23. An isolated nucleic acid molecule that encodes anantibody or antigen-binding fragment thereof that binds specifically tohuman B7RP1, wherein the antibody or antigen-binding fragment thereofcomprises any one of SEQ ID NOS: 44-58 and 70-76.
 24. The nucleic acidmolecule of claim 23, wherein the antibody or antigen-binding fragmentthereof is a single-chain antibody, a single-chain Fv antibody, an Fabantibody, an Fab′ antibody, or an (Fab′)₂ antibody.
 25. The nucleic acidmolecule of claim 23, wherein the antibody or antigen-binding fragmentthereof is a fully human antibody.
 26. The nucleic acid molecule ofclaim 23, wherein the antibody or antigen-binding fragment thereofblocks interaction between B7RP1 and ICOS.
 27. A vector comprising thenucleic acid molecule of claim
 23. 28. A host cell comprising the vectorof claim
 27. 29. The nucleic acid molecule of claim 1, 2, 3, 5, 13, 16,17, or 23, wherein the antibody or antigen-binding fragment thereofblocks T cell-dependent B cell antigen response.
 30. A method ofproducing an antibody or antigen-binding fragment thereof, that bindsspecifically to human B7RP1 comprising: a) culturing under appropriateconditions a host cell transformed or transfected with the nucleic acidof any of claim 1, 2, 3, 5, 13, 16, 17, or 23; and b) isolating theantibody or antigen-binding fragment thereof produced by the expressionof the nucleic acid.